DIFFERENTIATION OF THE C ELEGANS PHARYNX

DIFFERENTIATION OF THE C ELEGANS PHARYNX

3.7.         Differentiation of the C.elegans Pharynx

Pharynx in C.elegans is generated by two sets of cells. One group of pharyngeal precursors dependent on the maternal skn-1 gene and comes from the EMS cell. The second group of pharyngeal precursors comes from the ABa blastomere give the second group of pharyngeal precursors and it is dependent on EMS cell govern GLP-1 signalling, which activates the transcription factor-encoding pha-4 gene and that activates almost all of the pharynx- specific genes. The pha-4 transcription factor resembles the mammalian HNF3p protein.

3.8.         Vulva formation in C.elegans

C.elegans is a hermaphrodites nematode. C.elegans is male during the early development and at that time gonad produce sperm and store it for future use. They develop ovary during the later stage of development. Fertilization takes place inside the vulva when the egg gets roll by the region of sperm storage, After fertilization, fertilized egg get to pass down outside the body of a vulva.  In C.elegans formation of the vulva characterizes a case in which one inductive signal creates a variety of cell types. Vulva formation takes place during the larval stage, vulva form by six cells called the vulval precursor cells (VPCs), which give rise to 22 cells so that a mature vulva contain 22 cell form during postembryonic (larval) development.

During the L1 larval stage, eleven Pn.p cells (P1.p, P2.p, P3.p,..., P11.p) are formed as the posterior daughters of the Pn ventral neuro-ectoblasts.

Just before the end of the first larval stage, Wnt signalling takes place which induces expression of Hox gene lin-39 in six vulval precursor cells (6 Pn.p, from P3.p to P8.p become VPCs). The Lin 39 expression six vulva precursor cells are formed by vulval equivalence group. The lin 39 also maintains the VPCs as polarized epithelial cells. These six VPCs become competent to respond to an inductive signal or developmental signal that produced later during vulval development or become competent to Lin3 signalling.

3.8.1.     Hox genes and VPC competence group

At the establishment of the third larval stage, vulval precursor cells fates are determined by the collective action of the RTK/RAS/MAP kinase (LIN-3 epidermal growth factor (EGF) signalling) and the LIN-12 Notch signalling pathways. WNT signalling takes place via BAR-1 beta-catenin.

Within P3.p to P8.p expression of lin-39 takes place. Two main roles of lin-39, first, inhibits fusion with hypodermis via inhibition of EFF-1 expression.

(prevention of VPCs fusion) and second stimulating cell division, responsiveness to LIN-3. MAB-5 cause inhibition of lin-39 especially occurs in more posterior cells including P7.p and P8.p.

3.8.2.     Roles of LIN-39

Anchor cell, which is present within gonad, responsible for differentiation of hypodermis cell to vulva precursor cells (VPCs). The anchor cell is specified from two somatic gonadal cells during the end of the L2 stage, from two somatic gonadal cells cause specification of anchor cell. The basement membranes get to break down by anchor cell which is secreted by the gonad and the epidermis and maintain the separation of these two tissues. the anchor cell extends the process to the center of the vulF cells a process get extends by anchor cell and forms a hole in the epidermis. Through the vulE cells, anchor cell can pass and locate the vulF cells.

Anchor cell carried out paracrine signalling and induces vulval differentiation by secreting Lin3 that activates the receptor tyrosine kinase LET-23 (EGF receptor).

A concentration gradient gets the form by LIN-3 protein. The highest concentration of LIN-3 protein is received by the primary VPC closest to the anchor cell and generates the central vulval cells. The two VPCs adjacent to it (P5.p and P7.p) receive a lower amount of LIN-3 and become the lateral vulval cells.  VPCs farther away from the anchor cell become hypodermis because it does not receive enough LIN-3 from anchor cell.

When lin3-Let23 interaction on P6.p takes place. The RTK/RAS/MAP kinase signal transduction pathway carried out. specifies the primary (1°) vulval fate, firstly it activate Let60 (Ras) then it activates Lin 45(Raf), then it activates MEK-2 and MAK-1. The activation of protein block eff-1 block cell fusion. 

MAPK pathway target two transcription factors LIN-3l and LIN-1. LIN-3l and LIN-1 both get phosphorylated by MAP kinase MPK-1. Multivulva phenotype resulted from Inactivation of either lin-1or lin-31. The complex of  LIN-1and LIN-31 disrupted by LIN-31 phosphorylation. Vulvaless phenotype resulted from non-phosphorylated protein. lin-31 mutants during the L3 stage display both vulvaless and multivulva phenotypes Inappropriate divisions of Pn.p cells during the L2 stage display by  LIN-31, suggesting also a role in the proper specification of VPCs.

P6.p mediated lateral signalling, in response to induction by AC signal, as a result, LIN-12 Notch signalling ( juxtacrine signalling ) takes place by the release of Lin 12 ligand, which get bind to its neighbours P5.p and P7.p in turn cause inhibits the expression of the 1° fate (a process termed lateral inhibition) firstly and secondly mediates the secondary (2°) fate in P5.p and P7.p. MicroRNA, mir-61get activated by Notch signal, as a result cause repression of gene that would specify central vulval fate.

Tertiary (3°) fate get adopted by the VPCs (P4.p, P3.p and P9.p ) which receive neither inductive nor lateral signals, in turn, they fuse with the epidermis (hyp7) after undergoing one round of cell divisions.

Model of VPC pattern formation. 1°-2° patterning mechanism shown here within three of the six VPCs (P6.p, P7.p and P8.p). Anchor cell release Lin3 and LIN-3 acts in a graded fashion with P6.p receiving more signal than P7.p. LET-23 activation promotes the 1° fate, inhibits LIN-12 protein levels and cause the production of DSL ligands for LIN-12.  2° fate promoted by LIN-12 activation in turn cause inhibition of  response to LET-23 activation

3.8.3.     Mutational effect

VPCs will not form a vulva if the lin-3 gene is mutated and become part of the hypodermis (skin). If the anchor cell is destroyed, the three outer cells, which normally form hypodermis, generate vulval cells instead.

In the case of lateral signalling other VPCs, P7.p becomes 1° in a lin-15 multivulva mutant in the absence of the gonad. The two cells adopt a 1°-2° or 2°-1° pattern at random If P8.p is also present. P7.p

In the case of sequential signalling, mosaic analysis of let-23.  yellow, 3° fate; red, 2° fate; blue, 1° fate. The genotype of VPCs in the key mosaic animal as follows (-/- lacking let-23 activity) (+/+let-23activity present). all VPCs become 3° in the absence of let-23. In mosaic animals which have let-23 in P6.p but lack let-23 in P5.p and P7.p show that the vulva is usually wild-type. This observation indicates for presumptive 2°VPCs, let-23(+) is not required.

The pattern of VPC cell fates direct level of LIN-3. The six VPCs adopt the 3°-3°-2°-1°-2°-3° pattern in each wild-type case. Animals with reduced lin-3 function (lin-3(rf) show lin-3 reduction of function) display that P6.p induced to 1°. A highly penetrant vulvaless phenotype resulted from a further decrease in lin-3 function. Through anchor cell excessive lin-3  (lin-3(+ + +)) results in an expansion of the pattern, means more than one central cell.

In lin-17 mutants, vulva gets the form by  P5.p and P6.p or may be posterior pseudo-vulva get from by P7.p.

Constitutive 23 mutation causes a MUltiVulva (Muv) phenotype. Typically have a single functional vulva in multivulva hermaphrodites and additional ventral protrusions vulval tissue form each pseudovulva. A psuedovulva posterior to the normal vulva resulted from a mutation within the reversed polarity of P7.p 2° lineage.

There are fewer VPC divisions in the cye-1 mutant.


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