MESODERMAL ORGANS- Body wall muscle (Muscle Part II)

Nematode somatic muscle - Development of somatic muscle - Structure of somatic muscle - Muscle basal lamina - Innervation of somatic muscle - Cell list - Back to Contents

Nematode somatic (body wall) muscle

All nematode somatic muscles are longitudinal (Bird, A. F. and Bird, J., 1991). In C. elegans the 95 rhomboid-shaped body wall muscle cells are arranged as staggered pairs in four longitudinal bundles located in four quadrants (MusFIG28A-D and MusFIG29). Three of these bundles (DL, DR, VR) contain 24 cells each, whereas VL bundle contains 23 cells (this asymmetry appears to result from a gap on the ventral left quadrant of the embryo slightly posterior to the gonad primordiurn (Sulston J. E. and Horvitz H. R., 1977. See MusFIG32 below). Muscles are always separated from the underlying hypodermis and nervous tissue by a thin (approximately 20 nm) basal lamina. This basal lamina remains intact within the synaptic regions except for NMJs made in the nerve ring between the RIML/R motor neurons and their target muscle arms (See glrFIG5). The muscle arms innervated by RIML/R make small spurs which pierce through the basal lamina separating the muscle plate from the nerve ring to access these neurons.

A typical somatic muscle cell has three parts; the contractile filament lattice (spindle), a noncontractile body (muscle belly) containing the nucleus and the cytoplasm with mitochondria, and muscle arms, slender processes that extend to either ventral or dorsal nerve cords (See Part I)(MusFIG30). Somatic muscle nuclei are oblong (ovoid), intermediate in size between neuronal and hypodermal nuclei, and have a small, spherical nucleolus (MusFIG31). Viewing by DIC microscopy, their nucleoplasm appears granular in L1 (See Fig 6 in Sulston J. E. and Horvitz H. R., 1977) but becomes smooth in L2 and remains so throughout the rest of the development.

The development of somatic muscle

The body wall muscle cells are derived from D, C, AB and MS lineages. At hatching 81 of the 95 cells are present. 14 more muscle cells are generated postembryonically from the MS.apaapp lineage (Sulston J. E. and Horvitz H. R., 1977; Sulston J. E. et al, 1983). Of the 81 body muscles of the newly hatched larva, 80 are generated in symmetrical fashion from MS, C, and D lineages; 20 come from the D blast cell which exclusively generates body muscle cells, 16 come from Cp, 16 come from Ca, 5+1+3 come from Mspp, 6 come from MSpa, 5+3+1 come from MSap, 4 come from MSaa (MusFIG32. Numbers correspond to muscle cell numbers; (A)-(D) correspond to muscle quadrant rows referred to in MoW Table 2 as well as MusTable5 below)(Sulston J. E. et al, 1983). The remaining cell is generated by ABprpppppa and is one of a group of four muscles generated preanally by ABp(l/r)pppppa lineages (the other three cells become the anal depressor muscle, the sphincter muscle, and one of the two stomato-intestinal muscles).

Myoblasts are born after the end of gastrulation at about 290 min of embryonic development (MusTable1). At this stage muscle cells are seen in two lateral rows next to the seam cells and some muscle cells have not yet undergone their terminal divisions. During this time, hemidesmosome components start to accumulate in hypodermis in a diffuse fashion and similarly muscle cells start accumulating muscle components diffusely (MusFIG33A). Subsequently, around 350 min of development, the muscle cells migrate dorsally and ventrally to contact the ventral and dorsal hypodermis (Coutu Hresko M. et al 1994, 1999) (MusFIG33B). All muscle cells finish their divisions before assuming their final positions. Cell-cell contact induces the components of the muscle contractile apparatus to coalesce at the membrane near the contact points and fibrous organelle components (MH5 protein, intermediate filaments) become restricted to specific regions of the hypodermis adjacent to muscle. Basement membrane components are recruited to regions of contact between muscle cells (MusFIG33C). Hypodermal myotactin accumulates adjacent to where the contractile apparatus is forming in the muscle. By two-fold stage of development, muscle cells become flattened and myofilament lattice assembly begins with positional cues laid down in the basal lamina and muscle cell membrane( MusFIG33D)(Williams B.D. and Waterston R.H. 1994; Coutu Hresko M. et al 1994). Sarcomeres become organized into oblique striations and the interlocking arrangement of the rhomboid-shaped body wall muscle cells in separate bundles becomes apparent. At the earliest stage when individual muscle cells become discernable, each cell is two A-bands wide, in the adult, they may have grown to be as wide as ten A-bands, approximately 100 microns (Moerman D. G. and Fire A., 1997). During this time, myotactin remains adjacent to the forming contractile apparatus and its organization follows the oblique striations of the muscle. In contrast, components of fibrous organelles become organized in circumferentially oriented bands restricted to regions where hypodermis is adjacent to muscle. By three-fold stage (520 min after first cleavage of the embryo at 25°C), myotactin is seen to colocalize with fibrous organelle components in these bands (Coutu Hresko M. et al 1999).

Development at 25°C

MusTable 1: EMBRYONIC DEVELOPMENTAL EVENTS IN MUSCLE, HYPODERMIS AND BASEMENT MEMBRANE (Also see WA Handbook- Hypodermis)
<290 min Both dorsal and ventral hypodermal cells contain diffusely distributed myotactin, intermediate filaments (IF) and MH5 antigen. Dorsal hypodermal cells have not yet interdigitated.
290 min Muscle cells are born, and localized adjacent to seam cells on lateral sides. Vinculin, integrins and myosins are diffusely distributed in muscle cells (MusFIG33A).
310 min Muscle cells migrate towards dorsal and ventral hypodermis. Dorsal hypodermal cells have almost completed their interdigitation. Within the hypodermis, hemidesmosome components (IF and MH5 antigen) accumulate around the muscle cell contact regions .
350 min Muscle cells become organized into quadrants next to dorsal and ventral hypodermis. They are still round cells but become polarized; muscle components localize adjacent to muscle-muscle and muscle-hypodermis contact sites. Within basement membrane, myotactin and UNC-52/perlecan localize near muscle-muscle contact regions. Hemidesmosome components become restricted to regions of hypodermis adjacent to muscle cells. Dorsal hypodermal cells have completed their interdigitation (MusFIG33B).
420 min (1.5 fold stage) Muscle cells flatten. Myotactin and UNC-52/perlecan in basal lamina and IF and MH5 antigen in hypodermis colocalize with muscle components (MusFIG33C).
430 min Attachment structures form and lattice components organize into sarcomeres; UNC-52/perlecan and aPAT-2/bPAT-3/integrin become organized along basal muscle cell membrane in structures resembling cell-matrix adhesion complexes. DB and M-line components of muscle assemble at these sites. The thick filaments of the lattice then assemble with the M-line and the thin filaments with the DBs.
450 min (2-fold stage) IF and MH5 antigens localize into hemidesmosomes in hypodermis but myotactin in basement mebrane is still associated with muscle structures (MusFIG33D).
>520 min (three fold stage) Myotactin associates with hemidesmosomes.
References: Coutu Hresko M. et al 1994; Sulston J. E. et al, 1983; Rogalski T. M. et al 2001

The structure of somatic muscle

The body wall muscle of C. elegans, as in all other nematodes, is obliquely striated. Although the filaments themselves are oriented parallel to the longitudinal plane of the muscle cell, adjacent structural units (M lines and dense bodies (DB)) are offset from each other by more than a micron, rather than being in register as in vertebrate cross striated muscle (Bird, A. F. and Bird, J., 1991; Waterston R. H. 1988). Therefore, the observed A-I striations are at an angle of 5-7° to the longitudinal axes of the filaments and the muscle cell, in comparison to 90° in vertebrate cross-striated muscle (MusFIG34A-D. MusFIG34A shows surface view of myofilament structure and filament packing. Note that the distance between DBs (black dots) on the longitudinal axis is more than 10 microns (y), while in the tranverse plane they are only about 1 micron apart (x), for a ratio of about 10:1 (Waterston R. H. pers comm.); In MusFIG34B the surface view is laterally extended to better indicate the offsetting of contractile units. Thick filaments are shown as brown lines, thin filaments as black lines, DB as black dots and M lines as brown dots. MusFIG34C shows a cross section of the myofilament lattice. In the muscle cell the sheet of filaments lies inside the muscle membrane which is separated from the underlying hypodermis with a (20 nm) basal lamina. Cuticle, in turn, lies outside the hypodermis. MusFIG34D shows a 3-D rendering of myofilament lattice as well as structure of the sarcomere. Inset illustrates orientation and proportion of thick filaments and position of M lines) (Also compare MusFIG34A-D and MusFIG36 with MusFIG11 in Part I)). This oblique arrangement of the sarcomeres is suggested to create a more evenly distributed muscle force application over basal lamina and cuticle allowing for smooth bending of the body instead of kinking (Burr A. H. J. and Gans C., 1998).

MusTable 2 - DIFFERENCES BETWEEN C. elegans AND VERTEBRATE STRIATED MUSCLES
C. elegans striated muscle Vertebrate striated muscle (see Muscle Part I)
Striated muscle cells do not fuse to form a multinucleated muscle cell Muscle cells fuse to form mutinucleated myofiber (muscle cell)
Thick filaments contain paramyosin (an invertebrate protein) Thick filaments do not contain paramyosin
Obliquely striated (adjacent filament units are not in register) Cross striated (adjacent filament units are in register)
Thick filaments are 10 mm in length and taper in diameter from 33.4 nm centrally to 14 nm distally Thick filaments are 1.6 mm in length and uniform in diameter (12-14 nm)
Thin filaments are 6 mm in length and 8 nm in width Thin filaments are 1 mm in length, and similar to nematode thin filaments in width
The packing arrangement of filaments is 12:1 (thin:thick) (MusFIG35 & MusFIG36) The packing arrangement of filaments is 6:1 (thin:thick)
Thin filaments are attached to dense bodies Thin filaments are attached to Z lines
Transmission of tension occurs laterally through lateral attachments from the dense bodies to the cuticle Transmission of tension occurs longitudinally through the attachment plaques at the end of the muscle cells
There are no T-tubules (See Muscle Part I) T-tubules are part of the triad junctions where E-C coupling occur.
Ref: Moerman D. G. and Fire A., 1997; Waterston R. H. 1988; Bird, A. F. and Bird, J., 1991; Zengel J. M. and Epstein H. F., 1980.

In similarity to myotendinous junctions of vertebrate skeletal muscle, the ends of the C. elegans somatic muscle cells contain thin (actin) filament attachment plaques (the ends of the terminal half I bands where microfilaments are attached to the cytoplasmic surface of the plasma membrane) through which each of the muscle cells adheres tightly to adjacent muscle cells within one quadrant (MusFIG37)(Coutu Hresko M. et al 1994, Francis R. and Waterston R. H. 1991). Although this may allow for some tension to be transmitted longitudinally between cells (force b in MusFIG37), the bulk of the tension created by muscle contraction is transferred to the exoskeleton/cuticle through lateral attachments (DB/FO's) which are distributed along the entire length of the cell (force a in MusFIG37) (Francis G. R. and Waterston R. H. 1985; Woo W. M. et al, 2004).

The highly ordered series of lateral attachments are made between muscle, hypodermis and cuticle and consist of basal lamina components, fibrous organelles (FO) of hypodermis, and dense bodies (DB) and M-lines of muscle (MusFIG38, MusFIG39 and MusFIG40A-C)(Hahn B. S. and Labouesse M., 2001; Cox E. A. and Hardin J. 2001) . FO, which are also known as transepidermal attachments (Ding M. et al, 2004), are homologous to vertebrate hemidesmosomes (HD) that anchor the intermediate filament network to the plasma membrane and basal lamina. Like HD, they are seen as two electron-dense plaques, one on each side of hypodermal membrane, which are connected by intermediate filaments that extend across the thin hypodermis and attach to the cuticle and basal lamina. FO's are restricted to the hypodermal regions that overlie muscle cells and they form concurrently with muscle development. In early embryonic stages, they are localized into longitudinal strips, however, during elongation of the embryo and as circumferential actin bundles form in hypodermal cells, they change into a circumferential stripe pattern. This pattern continues through the larval and adult stages (Ding M. et al, 2004).
The myofilament lattice is anchored to the muscle cell membrane and adjacent basal lamina by integrin-containing DB and M-lines. These, in turn, are linked to intermediate filament arrays which span the hypodermis and attach to the outside cuticle (Waterston R. H. 1988; Francis R. and Waterston R. H. 1991; Moerman D. G. and Fire A., 1997; Coutu Hresko M. et al 1999). DB and M-lines contain many of the components found in focal adhesion plaques including talin, and UNC-97/PINCH, besides integrins (MusTable3). Dense bodies also share some antigens with attachment plaques (Francis G. R. and Waterston R. H. 1985).
Loss of function in attachment structure components of DB and M-lines frequently results in detachment of body wall muscles from the cuticle supporting the hypothesis that these attachment structures function to promote mechanical strength between the muscle and hypodermis (MusTable3).

MusTable 3: MUSCLE ELEMENTS

THICK FILAMENT

encoding genes tissue/location structure function
Myosin Heavy Chain-A (mhcA) myo-3 Striated and nonpharyngeal nonstriated muscle Preferentially forms homodimers Recognized by mAb DM5.6 (Moorthy S. et al, 2000). Forms the central 1.8 mm of 10.6 mm long thick filament. MHC-A null animals have severely impaired thick filament assembly and die as embryos (Waterston R. H., 1989)
Myosin Heavy Chain-B (mhcB) unc-54(myo-4) Striated and nonpharyngeal nonstriated muscle Preferentially forms homodimers Predominantly outside the central area, forms the 4.4 mm arms on either side. MHC-B null animals are viable with weakly contractile muscles (Miller D. M. III et al 1983, Epstein H. F., 1986)
Myosin Heavy Chain-C (mhcC) myo-2 Pharyngeal muscle   (Ardizzi J. P., Epstein H. F., 1987)
Myosin Heavy Chain-D (mhcD) myo-1(let-75) Pharyngeal muscle   (Ardizzi J. P., Epstein H. F., 1987)
Myosin Light Chain

mlc-1

mlc-2

mlc-4

mlc-1, 2: body-wall, pharyngeal and possibly vulval muscles. Not seen in intestinal, uterine, anal depressor, and sphincter muscles

 

mlc-4: uterine muscle, intestinal muscle, gonadal sheath. Not seen in body wall muscle, vulval muscle, pharyngeal muscle (Shelton CA et al, JCB 1999).

 

mlc-1(null) mutants are wild type, whereas 90% of mlc-2(null) hermaphrodites arrest as L1 larvae due to pharyngeal muscle defects (this effect is sex specific, since mlc-2(null) males are essentially wild type). Functions of mlc-1 are redundant to those of mlc-2 in both body-wall and pharyngeal muscle. Function of mlc-2 is redundant to mlc-1 in body-wall muscle, but not in the pharynx. mlc-1(null) mlc-2(null) double mutants arrest as incompletely elongated L1 larvae, having both pharyngeal and body-wall muscle defects (Rushforth AM et al, 1998).

Paramyosin unc-15 Thick filaments a-helical coiled coil formed by 2 identical subunits

An invertebrate protein. Forms the central region of the thick filament. Myosin molecules assemble around paramyosin core. UNC-15 physically interacts with MHC-A. In unc-15 null animals normal thick filaments are lacking.

(Waterston R. H. 1988; Kagawa H. et al, 1989; Hoppe P. E. and Waterston R. H., 2000)

Twitchin unc-22 Located in the thick filament containing A-bands A large protein that belongs to Ig superfamily. Functions both in regulation of the actomyosin contraction-relaxation cycle, and in the final stages of sarcomere assembly. In vitro, UNC-22 can phosphorylate myosin light-chain peptides and can undergo autophosphorylation (Benian G. M. et al 1989, 1996)
THIN FILAMENT        
Actin act-1,2,3,4,5 All muscles Double stranded F-actin

Slide past the thick filaments during contraction.

(Krause M and Hirsh D. 1984; Krause M. et al, 1989)

Troponin

tnt-1 (mup-2)

tni-1; unc-27 (unc-99, tni-2); tni-3; tni-4

pat-10 (tnc-1); tnc-2

 

tni-1 ( body wall muscles); tni-2 (body wall muscles); tni-3 (body wall muscles); tni-4 (pharyngeal muscle) (Razia R. et al, 2001).

 

pat-10 (body wall, vulval and anal muscles).

3 subunits:

T (TNT-1)


I (TNI-1; TNI-2; TNI-3: TNI-4)


C (TNC-1; TNC-2)

Troponin complex regulates contraction in response to Ca++ .

Troponin T functions in inhibition of thin filaments. In troponin T mutants thin filaments are unregulated and generate aberrant force production. mup-2 (null) mutants are defective for embryonic body wall muscle cell contraction, sarcomere organization, and muscle cell positioning and die as kinked larvae (Myers C. D. et al, 1996; Allen T. S. et al, 1995)

Troponin I plays an essential role in interaction with troponin C and is also necessary for muscle filament assembly and motility during development. In unc-27 mutants dense body positioning is impaired and sarcomeric structure is less well defined, including small islands of thin filaments interspersed within the overlap region of A bands and even within the H zone (Burkeen A. K. et al 2004).

Troponin C is the calcium-binding component of the troponin complex of actin (thin) filaments. Tropomyosin and troponin C mutations prevent activation (disinhibition) of thin filaments and thus block muscular contraction.
PAT-10 is essential for muscle contraction and therefore for completion of embryonic morphogenesis and elongation. pat-10 mutant animals arrest at two-fold stage (Terami H. et al, 1999) .

Tropomyosin lev-11(tmy-1) Body wall muscles, anal muscles, vulval muscles, pharyngeal muscles and the male sex muscles a-helical coiled coil Each tropomyosin molecule has seven evenly spaced regions with similar amino acid sequences, each of which is thought to bind to an actin subunit in the filament. Tropomyosin blocks the myosin binding sites on actin in resting state. Ca2+ (binding to troponin) is thought to relieve the tropomyosin blockage of the interaction between actin and the myosin head (Alberts B. et al (ed.s) Molecular Biology of the Cell, 2002; Kagawa H. et al, 1995, 1997; Anyanful A. et al, 2001).
Cofilin unc-60A/B     UNC-60A depolymerizes actin filaments and inhibits actin polymerization, whereas UNC-60B strongly binds to F-actin without depolymerizing it and, through binding to G-actin, changes the rate of actin polymerization depending on the UNC-60B:actin ratio. Both proteins are implicated in the regulation of actin filament assembly in body wall muscle (Ono S. et al, 1999; Ono S. and Benian G. M., 1998).
UNC-78 unc-78 Body wall muscle Actin interacting protein 1 homologue. A WD-repeat protein. Enhances actin filament disassembly in the presence of actin depolymerizing factor (ADF)/cofilin. In mutant animals organization of actin filaments is disrupted and large actin aggregates are formed in body wall muscle cells (Ono S. 2001; Mohri K and Ono S., 2003)
UNC-87 unc-87 All muscles (Goetinck S. and Waterston R. H., 1994a) The molecular domain structure of UNC-87 reveals similarity to the C-terminal repeat region of the smooth muscle actin-binding protein calponin, but there are no obvious orthologs outside of nematodes. The protein is localized in the I-band of bodywall muscle and associates with actin stress fibers. It may function as an actin-bundling protein. unc-87 mutants show degeneration of myofilaments and deterioration of the muscle as a result of contraction and mechanical stress. These mutants are severly immobile and also have egg-laying defects (Goetinck S. and Waterston R. H., 1994a, 1994b; Kranewitter W. J. et al, 2001).
ATTACHMENT COMPLEXES        
MUSCLE COMPONENTS       Note that vinculin and a-actinin are localized to DBs only and UNC-89 is localized to M-lines only (Rogalski T. M. et al 2000).
Vinculin deb-1(pat-8)     Localized to base of DBs and attachment plaques and is required for attaching actin and myosin filaments to the sarcolemma (Francis G. R. and Waterston R. H. 1985; Barstead R. J. and Waterston R. H. 1989). deb-1 mutant animals lacking vinculin arrest development as L1 larvae. In these animals, embryonic elongation is interrupted at the twofold length, so that the mutants are shorter than wild type animals at the same stage. The mutants are paralyzed and has disorganized muscle (Barstead R. J. and Waterston R. H., 1991).
UNC-95 unc-95   LIM domain protein UNC-95 is required for the proper assembly of muscle attachment sites. The initial deposition of UNC-52/Perlecan in the basal lamina and PAT-3 accumulation in the basal sarcolemma occurs normally in unc-95 mutant embryos. However, the recruitment of DEB-1/vinculin to these nascent attachments sites requires UNC-95. Thus, UNC-95 is not necessary for the initiation of the assembly complex, but for downstream processes of sarcomere organization. In mutant animals both the I and A bands are disorganized. This suggests that UNC-95 has a role in both the DB and M-line assembly pathways (Broday L. et al, 2004)
PINCH unc-97 Body wall muscle, sex muscles, touch neurons A conserved LIM protein. UNC-97 colocalizes with b-integrin to DBs and M lines. unc-97 mutants have shallow, disorganized sarcomeres (Hobert O. et al. 1999; Zengel J. M. and Epstein H. F., 1980).
UNC-98 unc-98 Body wall muscle Zinc finger protein. UNC-98 resides at muscle cell nuclei, M-lines, and possibly at DB. It interacts directly with UNC-9. unc-98 mutants have disorganized myofilament lattices with fewer or less distinct A and I bands (Mercer K. B. et al, 2003)
a-actinin atn-1 Body wall muscle, pharynx   Ca2+-binding actin-bundling protein. The principal cross-linker of actin filaments which stabilizes and anchors thin filament arrays. Localized throughout the DB (Francis G. R. and Waterston R. H. 1985 ; Barstead R. J. et al. 1991).
Talin       Localized to DBs and M lines. b-integrin but not vinculin is required for organization of talin at the muscle membrane (Moulder G. L. et al. 1996).
ILK pat-4 Bodywall muscle. Colocalizes with the integrin PAT-3 subunit Serine/threonine kinase PAT-4 is an integrin-linked kinase and also functions as an adaptor protein within integrin adhesion complexes. In pat-4 (null) mutants, embryonic muscle cells form integrin foci, but the subsequent recruitment of vinculin and UNC-89 as well as actin and myosin filaments to these in vivo focal adhesion analogs is blocked. Conversely, PAT-4/ILK requires UNC-52/perlecan, the transmembrane protein integrin, and UNC-112 to be properly recruited to nascent attachments (Mackinnon A. C. et al, 2002; Labouesse M and Georges-Labouesse, 2003)
Actopaxin pat-6     Required to recruit UNC-89 and myofilaments to newly forming attachments, as well as to reposition these attachments so that they form the highly ordered array of DB and M line attachments that are characteristic of mature muscle cells . PAT-6 is not required for the deposition of UNC-52/perlecan in the basal lamina, nor for the initiation of attachment assembly, including the clustering of integrin into foci and the recruitment of attachment proteins PAT-4/ILK, UNC-112 , and vinculin from the cytosol (Lin X. et al, 2003; Labouesse M and Georges-Labouesse, 2003).
UNC-89 unc-89   A large modular protein composed of Ig and signal transduction domains. Localized to M-lines (Benian G. M. et al. 1996). unc-89 mutants have disorganized muscle structure in which thick filaments are not organized into A-bands, and there are no M-lines (Waterston, R.H., et al 1980).
DIM-1 dim-1 Localizes to the region of the muscle cell membrane around and between the DB Contains three Ig repeats In dim-1 mutants muscle structure is slightly disorganized, and the myofilament lattice is not as strongly anchored to the muscle cell membrane as it is in wild-type muscle. DIM-1 may play a role in stabilizing the thin filament components of the sarcomere upon muscle tension (Rogalski M. et al, 2003)
UNC-112 unc-112 Colocalizes with PAT-3/b-integrin in both adult and embryonic body wall muscle. FERM domain protein. A membrane-associated, intracellular protein that is required for the proper spatial localization of PAT-3/b-integrin in the basal cell membrane. UNC-112 is not required to organize UNC-52/perlecan in the basal lamina nor for DEB-1/vinculin to associate with PAT-3/b-integrin (Rogalski T. M. et al 2000). In the absence of the UNC-112 protein, actin and myosin filaments do not attach to the muscle cell membrane and the animals arrest at the two-fold stage (Williams B. D. and Waterston R. H.,1994).
BASAL LAMINA (BASEMENT MEMBRANE) COMPONENTS        
Perlecan unc-52 Basal lamina between the bodywall muscle cells and the hypodermis. UNC-52/Perlecan is concentrated at the DB and M line sites. Extracellular matrix core heparan sulphate proteoglycan Major heparan sulphate proteoglycan of the extracellular matrix. UNC-52 is expressed in muscle cells and assembles into the basal laminae immediately adjacent to the cells that synthesize it (Moerman D. G. et al, 1996). UNC-52 is necessary for the proper localization of PAT-3/b-integrin to the muscle cell membrane and anchoring dense bodies and M-lines. In unc-52 null animals myofilament lattice can not assemble during late embryogenesis resulting in a paralysed embryo with arrested elongation at two-fold stage and terminal (Pat) phenotype (Williams B. D. and Waterston R. H. 1994; Rogalski T. M. et al 1995, 2001, 1993).
a-integrin pat-2 Transmembrane proteins found at the cell-matrix adhesion sites of both DBs and M-lines of body wall muscles. Also found in vulval, uterine, anal depressor, and anal sphincter muscle attachments, as well as within the myoepithelial sheath cells of the gonad. Colocalizes with PAT-3. Forms heterodimers with b-integrin subunit a-PAT-2/b-PAT-3 heterodimers are transmembrane components of DBs and M-lines, the structures that anchor, respectively, the actin and myosin filaments to the cell membrane. In embryos homozygous for mutations in the pat-2 gene, myosin and actin are not organized into sarcomeres in the body wall muscle cells, and DB and M-line components fail to assemble. These animals arrest at two-fold stage (Williams B. D. and Waterston R. H. 1996; 1997).

b-integrin

pat-3 Transmembrane proteins found at the cell-matrix adhesion sites of both DBs and M-lines of body wall muscles. Also found in anal depressor muscle, sphincter muscle, stomatointestinal muscles, vulval and uterine muscles. Colocalizes with PAT-2. Not detected in pharyngeal muscle. Forms heterodimers with a-integrin subunit Recognized by mAb MH25. a-PAT-2/b-PAT-3 heterodimers are transmembrane components of DB and M-lines, the structures that anchor, respectively, the actin and myosin filaments to the cell membrane. PAT-3/b-integrin is required for DEB-1/vinculin to localize to the base of the dense bodies. In embryos homozygous for mutations in the pat-3 gene, myosin and actin are not organized into sarcomeres in the body wall muscle cells, and DB and M-line components fail to assemble (Gettner S. N. et al, 1995 ; Williams B. D. and Waterston R. H. 1996; 1997)
Laminin

epi-1 (a)

lam-1 (b)

lam-2 (g)

lam-3 (a)

    Laminin organizes into a polygonal array on the surface of body wall muscle cells. In a subunit mutants muscle cells may fail to properly adhere to the overlying hypodermis. In these animals muscle adhesion complexes and myofibrillar components are improperly positioned and the cytoskeleton is defective in the hypodermis adjacent to where the muscle cells attach. This suggests that laminin plays a crucial role in organizing a supramolecular architecture comprising extracellular matrix, receptors and cytoskeletal components (Huang C. et al, 2003).
Nidogen nid-1 NID-1 is concentrated under dense bodies, at the edges of muscle quadrants, and on the sublateral nerves that run under muscles.  

Although nidogen can form a ternary complex with type IV collagen and laminin, C. elegans nidogen homologue NID-1 is not essential for type IV collagen assembly into basal laminae and nid-1 mutant animals have no overt morphological or behavioral phenotypes except decreased fecundity and some nerve positioning defects (Kang S. H. and Kramer J. M. 2000; Kim and Wadsworth, 2000).

SPARC ost-1 Body wall muscle, sex muscles   Localized immediately adjacent to body wall muscle and is concentrated at the boundaries between muscle cells similar to type IV collagen (Fitzgerald M. C. and Schwarzbauer J. E., 1998)
Type IV collagen emb-9 (a1-like); let-2 (a2-like) Expressed primarily in mesoderm; body wall muscles, presumptive GLR cells, HMC, coelomocytes, enteric muscles, distal tip cells, spermatheca, vulval and uterine muscles Type IV collagen has a triple helical structure. In basal laminae, type IV collagen molecules form a complex polygonal network stabilized by intermolecular disulfide bonding. In emb-9 and let-2 mutant embryos body wall muscle quadrants are disrupted with gaps. Since the first appearance of these gaps correspond to the beginning of muscle contractile activity in the embryo, it is suggested that the muscle cells detach from the body wall during contractions in these mutants (Gupta M. C., et al 1997; Graham P. L., et al 1997).
HYPODERMIS COMPONENTS        
Myotactin let-805 Colocalizes with hypodermal intermediate filaments and fibrous organelles 498 kD hypodermal transmembrane protein Recognized by mAb MH46 . Myotactin helps maintain the association between the muscle contractile apparatus and FO's. let-805 mutant animals, although they begin to twich at the 1.75-fold stage, never roll within the egg shell and fail to elongate beyond the two-fold stage. Muscle cells detach from the body wall in these embryos (Coutu Hresko M. et al 1999).
MUP-4 mup-4 Colocalizes with hemidesmosome structures overlying the body wall muscle and other mechanical attachment sites of hypodermis (e.g. touch neuron channels). Hypodermal transmembrane protein Functions in intermediate filament tethering, hypodermis - extracellular matrix adherence and transmission of mechanical forces to the cuticle. MUP-4 is essential in embryonic hypodermal morphogenesis and maintenance of muscle position. In mup-4 mutant animals muscles detach from the body wall (Hong L. et al, 2001).
MUA-3 mua-3 Localizes to hypodermal hemidesmosomes at the sites of skeletal muscle contact, other muscle-epithelium attachment sites (vulval muscles, the anal depressor muscle, the anal sphincter muscle), touch neuron channels, amphid, phasmid, and IL sensillar openings, and excretory duct and pore cells. Not localized at the uterine seam or in the pharynx. Hypodermal transmembrane protein Links hypodermal/epithelial intermediate filaments to the cuticle at the sites of mechanical stress and functions to transmit muscle force across the hypodermis to the cuticle. In mua-3 mutant animals muscles detach from the body wall (Bercher M. et al, 2001) .
VAB-19 vab-19 Hypodermis. Colocalizes with intermediate filaments. Ankyrin repeat protein vab-19 mutants are defective in muscle attachment to the hypodermis. Though hypodermal attachment structures form normally in these animals, they do not remain localized to muscle-adjacent regions. VAB-19 is , therefore, not required for the assembly of attachment structures but is required for their localization to muscle-adjacent regions of hypodermis. These mutants also display aberrant actin organization in the hypodermis. VAB-19 localization requires myotactin (Ding M. et al, 2003).
Spektraplakin vab-10A Hypodermis, pharynx and along mechanosensory axons. Colocalizes with myotactin in adults. Ortholog of vertebrate HD component plectin. Loss of VAB-10A impairs the integrity of FOs leading to detachment of hypodermis from the cuticle and muscles (Bosher J. M. et al, 2003)
MH5 antigen   Localizes to hypodermal FO's in body wall. Also localizes in touch neuron channels, tonofilament anchoring desmosomes in pharyngeal marginal cells, vulval muscle-hypodermis, uterine muscle-hypodermis and anal depressor muscle-hypodermis attachment sites. 300kD hemidesmosomal component. Recognized by mAb MH5 (Francis R. & Waterston R. H.,1985)
Intermediate Filaments ifa-2 (mua-6); ifa-3; ifb-1A/B Hypodermis (ifa-2, ifa-3, ifb-1A/B), pharyngeal marginal cells (ifb-1A/B), excretory cell and duct (ifb-1A/B), uterine epithelium (ifb-1B) All contain a central rod domain that forms coiled coils and determines IF assembly properties. Head and tail domains influence interactions among IF subunits during polymerization. IFA-3 forms heteropolymeric intermediate filaments in vitro with IFB-1.

Components of transdermal attachment structures (ifa-3 and ifb-1 embryonic; ifa-2 larval). mAb MH4 recognizes an epitope common to IFA-1, IFA-2, IFA-3 but not IFB-1.

Reduction of IFB-1 function by RNAi results in muscle detachment and disorganized myofilament lattice in embryos.

IFA-2 is required for larval muscle attachment ; ifa-2 mutants display muscle detachment, progressive paralysis and arrest during the L1 or L2 stage.

Reduction of IFA-3 function by RNAi results in embryonic lethal phenotype with arrest during embryonic elongation and muscle detachment.

(Francis R. & Waterston R. H.,1985; Plenefisch J. D. et al, 2000; Hapiak V. et al, 2003; Karabinos A. et al, 2001, 2002, 2003; Woo W. M. et al, 2004)

Muscle basal lamina (basement membrane)

As in other organisms, basal laminae are thin sheets of specialized extracellular matrix that contains type IV collagen, laminin, nidogen, SPARC, and perlecan in C. elegans. Somatic muscle quadrants in the body run inside tubes of basal lamina which separates them from the pseudocoelomic cavity and underlying hypodermis and nervous tissue (MusFIG41 panel B. Note this section roughly corresponds to Slidable worm section #415). The neuronal processes that run from the ventral side to the dorsal side (commissures) run under this basal lamina and between muscle and hypodermis. In the head, the basal lamina is extended around the muscle arm plate and separates the muscle arms from the nerve ring (see MusFIG3 in Part I). This extension of basal lamina terminates onto the cylinder of sheetlike processes of the GLR cells anterior to the nerve ring. Similar to the body muscles, the head muscle quadrants run inside tubes of basal lamina in more anterior regions of the head (MusFIG41 panel A. Note this section roughly corresponds to Slidable worm section #50).

Innervation of somatic muscle

The characteristic undulatory movement of nematodes is generated by alternate contraction of the dorsal and ventral longitudinal muscle rows creating sinusoidal waves in the dorso-ventral plane (the animal lies on its lateral side on the substrate). As the dorsal muscles are activated, the ventral ones are reciprocally inhibited by inhibitory GABAergic motor neurons and vice versa. The bending against the substrate results in forward locomotion from backward waves. Depending on the basis of their synaptic input the somatic muscles fall into 3 groups; the anteriormost four somatic muscle cells in each quadrant (head muscles) are innervated by motor neurons in the nerve ring (refer to MusTable 4 and MusTable5 below and see originals in White J.G. et al., 1986- table 2 and 3), the next four cells in each quadrant (neck muscles) receive dual innervation from motor neurons of the nerve ring and the ventral nerve cord and the remainder (body muscles) are exclusively innervated by ventral cord motor neurons (MusTable4, MusFig32)(White J.G. et al., 1986; Bird, A. F. and Bird, J., 1991). The muscle cells in each row of a muscle quadrant are electrically coupled to their neighbors through gap junctions. Also, the muscle arms from the neck muscles make extensive gap junctions with the head mesodermal cell which may provide electrical coupling between the dorsal and ventral muscles in this region (White J., 1988).

Body. The members of each class of body motor neurons are evenly distributed along the length of the ventral cord and create a longitudinal, synaptic fate map of body muscles. These neurons include the cholinergic, stimulatory A and B type motor neurons (VA, VB, DA, DB, AS) and inhibitory, GABAergic D type motor neurons (VD, DD). The cell bodies of all of these neurons are located in the ventral nerve cord. It should be noted that the VC motor neuron class of egg-laying circuitry also innervate ventral body muscles. Unlike the head motor neurons (see below), the ventral cord motor neurons synapse onto either both dorsal or both ventral muscle quadrants thereby restricting the body's flexures in the dorso-ventral plane. The ventral muscles are innervated by VA, VB, VD and VC classes and dorsal ones are innervated by DA, DB, DD and AS neurons. At hatching, DA, DB, and DD are the only classes of motor neuron present in the ventral cord, the other classes are born postembryonically. An additional excitatory class of neurons, SABVL/VR/D, innervate anterior ventral body muscles in the L1 stage.
D type motor neurons in the body have processes that are postsynaptic corecipients at the dyadic NMJs of stimulatory motor neurons. Their synaptic inputs are onto diametrically opposite muscles where muscle is the only recipient of their synaptic signal. When a ventral or dorsal muscle group is activated by a cholinergic motor neuron, the opposite group of muscles are inhibited through the function of these GABAergic D type neurons. The two subclasses of these neurons (ventral D and Dorsal D) work as reciprocal pairs and cross inhibitors covering similar muscle sections such that where one is postsynaptic, the other is presynaptic and vice versa (on the dorsal side VDs are exclusively postsynaptic, receiving input from DA, DB, AS neurons as corecipients at NMJs. On the ventral side, VD neurons are presynaptic to body wall muscle. DDs are exclusively postsynaptic on the ventral side where they receive input from VA, VB and VC neurons as corecipients at NMJs. DD's are predominantly presynaptic to body wall muscle on the dorsal side). This arrangement allows for sequential cross-inhibition of the muscles during undulatory body movement. It must be noted that only DD neurons are present at hatching when they make NMJs on the ventral side and are postsynaptic to DA and DB motor neurons on the dorsal side. As the animal goes through the first larval stage DD neurons synaptically rewire and reverse their polarity and VD neurons are born establishing the adult type wiring pattern for this class of neuron (White J. G. et al 1978).
Additional nerve-muscle contacts occur along the length of the sublateral cords (Duerr J., Hall D. H. and Rand J., unpublished data). Most axons in these nerve cords show periodic swellings filled with synaptic vesicles and often have small presynaptic densities. The postsynaptic targets of these syaptic release zones are possibly the body muscles.

Head and Neck. The head of the animal is capable of making lateral movements as well as dorso-ventral flexures, especially during foraging behavior. It is postulated that lateral movements of the head become possible since synapses from each nerve ring motor neuron is restricted to two adjacent rows of cells as opposed to two complete quadrants. These synapses can also be made to adjacent rows of cells in different quadrants allowing for differential activation of muscles in adjacent bands and even in adjacent rows in one quadrant (White J.G. et al., 1986). Head motor neuron classes with fourfold symmetry (RME, SMB, URA) innervate all eight rows of muscle with no overlap while the motor neurons with sixfold symmetry (IL1) have regions of innervation that overlap with each other by one row on each side but not across the dorso-ventral midline. Motor neurons with bilateral symmetry (RIM, RMF, RMG, RMH) mostly innervate only the lateral four rows of head muscles. RIVL/R only innervate cells in the ventral rows as an exception to this (White J. G. et al., 1986, see table 2). RMD and SMD motor neurons are suggested to be the cross-inhibitors in the nerve ring though the pattern of cross inhibition is probably more complex in the head compared to the body. SMDs are postsynaptic to GABAergic RME neurons which themselves are postsynaptic corecipients at NMJs, and both classes of putative cross-inhibitory motor neurons receive extensive synaptic input from interneurons unlike D type body neurons which are only postsynaptic to ventral cord motor neurons at NMJs. The major source of synaptic input to RMD and SMD neurons comes from RIA interneurons which themselves receive prominent inputs from RIB interneurons.

Cell list - Numbers in parentheses correspond to the cell numbers (as shown in MusFIG32)

1. Somatic muscles

Ventral L quadrant

(1) MS.apapap
(2) MS.appapp
(3) MS.apppaa
(4) D.aapa
(5) D.aaaa
(6) D.aapp
(7) D.apaa
(8) MS.aapppaa
(9) C.apaaaa
(10) MS.aapppap
(11) C.apaaap
(12) MS.aappppa
(13) C.apaapa
(14) MS.aappppp
(15) C.apaapp
(16) MS.pappaa(l)
(17) M.vlap (postembryonically born)
(18) M.vlaa (postembryonically born)
(19) M.vlpa (postembryonically born)
(20) C.apappa
(21) M.vlpp (postembryonically born)
(22) C.apappp
(23) C.appppv

Ventral R quadrant

(1) MS.ppapap
(2) MS.pppapp
(3) MS.ppppaa
(4) D.papa
(5) D.paaa
(6) D.papp
(7) D.ppaa
(8) MS.papppaa
(9) C.ppaaaa
(10) MS.papppap
(11) C.ppaaap
(12) MS.pappppa
(13) C.ppaapa
(14) MS.pappppp
(15) C.ppaapp
(16) MS.pappa(r)
(17) M.vrap (postembryonically born)
(18) M.vraa (postembryonically born)
(19) C.ppappa
(20) AB.prpppppaa
(21) M.vrpa (postembryonically born)
(22) C.ppappp
(23) M.vrpp (postembryonically born)
(24) C.pppppv

Dorsal L quadrant

(1) MS.apaaap
(2) MS.apappa
(3) MS.apappp
(4) MS.appppa
(5) MS.apppap
(6) MS.appppp
(7) D.aaap
(8) D.appaa
(9) D.apap
(10) D.appap
(11) C.appaaa
(12) D.apppa
(13) C.apapaa
(14) D.apppp
(15) M.dlap (postembryonically born)
(16) M.dlaa (postembryonically born)
(17) C.apapap
(18) C.appaap
(19) C.appapa
(20) C.apppaa
(21) M.dlpp (postembryonically born)
(22) C.apppap
(23) C.appapp
(24) C.appppd

Dorsal R quadrant

(1) MS.ppaaap
(2) MS.ppappa
(3) MS.ppappp
(4) MS.pppppa
(5) MS.ppppap
(6) MS.pppppp
(7) D.paap
(8) D.pppaa
(9) D.ppap
(10) D.pppap
(11) C.pppaaa
(12) D.ppppa
(13) C.ppapaa
(14) D.ppppp
(15) M.drap (postembryonically born)
(16) M.draa (postembryonically born)
(17) C.ppapap
(18) C.pppaap
(19) C.pppapa
(20) C.ppppaa
(21) M.drpp (postembryonically born)
(22) C.ppppap
(23) C.pppapp
(24) C.pppppd


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