Unit 10 Axis Specification and Gastrulation in C elegans Things to look for during axis specification How does inherent polarity of the oocyte egg one cell zygote influence axis specification How are fates of founder cells and the primary germ layers established in the early embryo How do external cells become internalized during gastrulation What morphogenetic movements occur to shape the embryo during gastrulation Types of Morphogenetic Movements Invagination is the inward bending of a sheet of tissue example formation of the digestive tube archenteron Involution is the rolling of a sheet of tissue around an edge example cells moving around the lip of the blastopore Gilbert 10e Table 5 2 Types of Morphogenetic Movements Radial intercalation Gilbert 10e Table 5 2 Epiboly often occurs via radial intercalation of multiple cell layers Wolpert 2e Fig 8 26 Types of Morphogenetic Movements Ingression is the detachment of single cells from a sheet of tissue example primary mesenchyme in the sea urchin gastrula Gilbert 10e Table 5 2 Delamination involves the splitting of one sheet of tissue to form two Types of Morphogenetic Movements Convergent extension Convergent extension involves the directed rearrangement of cells to shorten a tissue along one axis convergence while lengthening the tissue along the orthogonal axis extension example marginal zone cells in the frog gastrula germ band in flies archenteron in sea urchins Wolpert 2e Fig 8 26 Blastomere Identity in C elegans Anterior Posterior progeny make pharynx and epidermis due to induction from MS makes mesoderm muscle pharynx makes gut C elegans blastomere specification and gastrulation Courtesy Tim Walston EMS daughters form two major germ layers mesoderm muscle and pharynx endoderm gut Step 1 Sperm entry point defines the posterior sperm normally enters at the posterior P granules post fertilization rearrangements polarize the zygote polar body normally marks the anterior Goldstein and Hird 1996 Development 122 1467 1474 Step 2 Early protein asymmetries established anterior and posterior domains in the cortex established AB sperm centrosome organizes microtubules for pronuclear migration PAR 3 6 P1 A and P domains establish where division will occur PAR 2 MEX 5 Adapted from Nance 2005 Bioessays 27 126 135 Cortical domains form after fertilization due to actin and myosin rearrangements A P Courtesy of Ed Munro PAR proteins polarize the zygote PAR 3 A P PAR 3 PAR 3 and PAR 6 are in the anterior Courtesy of Ed Munro PAR proteins polarize the zygote A P PAR 2 PAR 2 is in the posterior Gilbert 9e Fig 5 43 par proteins are required for polarity offspring of par 3 mutant mothers have symmetric P granule distribution courtesy Ken Kemphues Step 3 SKN 1 PIE 1 accumulate in P1 MEX 5 accumulates in AB AB P1 MEX 5 PIE 1 MEX 5 SKN 1 SKN 1 PIE 1 MEX 5 protein in AB leads to PIE 1 and SKN 1 degradation via the ubiquitin pathway in AB and later in ABa ABp and EMS for PIE 1 PIE 1 protein is in P1 AB PIE 1 GFP Courtesy Geraldine Seydoux Nomarski P1 MEX 5 and 6 are in the anterior A P movie MEX 5 GFP MEX 5 protein in AB leads to PIE 1 and SKN 1 degradation via the ubiquitin pathway in AB Courtesy of Ed Munro MEX 5 6 restrict SKN 1 protein expression to P1 SKN 1 shown wildtype mex 5 Schubert et al 2000 Mol Cell 5 671 682 Step 4 PIE 1 represses SKN 1 activity in P2 ABp ABa MEX 5 P2 PIE 1 SKN 1 EMS EMS PIE 1 SKN 1 MEX 5 protein in ABa ABp and EMS leads to PIE 1 degradation except in P2 PIE 1 counteracts SKN 1 in P2 SKN 1 is in EMS P2 2 cell 4 cell SKN 1 is in EMS and P2 but it only acts in EMS Gilbert 5e Fig 13 19 MEX 5 6 restrict SKN 1 PIE 1 protein to P1 wildtype mex 5 mex 6 RNAi SKN 1 P2 P2 EMS EMS PIE 1 P2 P2 PIE 1 prevents SKN 1 activity in P2 EMS Schubert et al 2000 Mol Cell 5 671 682 SKN 1 skin in excess is responsible for correct EMS fate wildtype Mutation in skn 1 results in EMS producing muscle and epidermis skn 1 mutant mother pharynx gut Gilbert 9e Fig 5 45 no pharynx no gut Step 5 SKN 1 activates med genes MED proteins MS mesoderm E endoderm MED proteins are transcription factors that activate transcription of mesodermal and endodermal genes in MS E respectively SKN 1 is needed to activate med gene transcription SKN 1 activates transcription of med 1 med 2 MED 1 GFP MS E wildtype Wild type skn 1 mutant mother Maduro et al 2001 Mol Cell 7 475 Blastomere Identity in C elegans A P progeny make pharynx due to induction from MS and epidermis makes mesoderm muscle pharynx What makes E different from MS makes gut ABp ABa MOM 5 Frizzled Step 6 A Wnt signal induces endoderm in E MS forms mesoderm P2 MOM 2 Wnt EMS The side of EMS touching P2 receives a Wnt signal When EMS divides the side in contact with P2 becomes the endodermal precursor E PIE 1 MED proteins MS mesoderm E P3 endoderm some progeny make germline mom 2 Wnt and mom 5 Fz are required for E specification WT Mom more mesoderm More mesoderm results because cells that normally form endoderm don t and become mesoderm instead Thorpe et al 1997 Cell 90 695 gut pharynx mom 2 mutant P2 induces E at the site where it makes contact remove P2 normal ABp ABp ABa EMS ABa P2 EMS move P2 ABp ABa EMS P2 gut granules form from cell that arises from EMS P2 contact adapted from work by Bob Goldstein Question What would happen for each of the following a normal P2 cell is combined with an EMS cell from an embryo derived from a mom 2 Wnt mutant mother a normal EMS cell is combined with a P2 cell from an embryo derived from a mom 2 Wnt mutant mother P2 specifies gut through MOM 2 Wnt WT P2 mom 2 EMS mom 2 P2 WT EMS Gut Thorpe et al 1997 Cell 90 695 Ea and Ep endodermal precursors undergo ingression Ea and Ep pop into the interior via ingression Putzke Rothman 2003 Curr Biol 13 R223 R225 Ea Ep undergo ingression Nance et al 2003 Development 130 5339 Apical constriction contributes to Ea Ep ingression Ea Ep P4 Lee Goldstein 2003 Development 130 307 Phosphomyosin II accumulates during apical constriction phospho myosin II Jen Lee Dan Marston Lee et al 2006 Curr Biol 16 1986 1997 Neighboring cells migrate to cover Ea and Ep MS and P4 extend protrusions that may cover the space left by Ea and Ep Putzke Rothman 2003 Curr Biol 13 R223 R225 P4 and MS cells actively extend over ingressing cells Nance Priess 2002 Development 129 387 Other cells ingress later and are covered by epidermis Mesodermal and germ cell precursors ingress after Ea and Ep The epidermis covers interior cells via epiboly ventral enclosure