HEDGEHOG SIGNALLING IN DROSOPHILA

HEDGEHOG SIGNALLING IN DROSOPHILA

4.5.         Hedgehog signalling:

In absence of hedgehog: in the absence of hedgehog its receptor patch binds to smoothened get become inactive and the ci activator of hedgehog signalling bind to PKA, Fu, Sufu, gsk-1, ck1, cos and SCF, thus get degraded and convert in to ci repressor, go to the nucleus, binds to the response element of hedgehog-responsive gene and repress the transcription of  hedgehog-responsive gene  (wingless).

In the presence of hedgehog: hedgehog binds to its patch receptor, so that smoothened not able to block the patch then signal transduction occur and ci receptor remain free to go to the nucleus and activate the transcription of hedgehog-responsive gene, thus wingless also get express and wingless protein secreted out of the cell through co-translation transport and move to its receptor present at anterior end of the compartment.

4.6.         Homeotic genes: Responsibility of the homeotic gene to assign a unique identifier to each segment and thus patterning along the anterior-posterior (AP) axis. Mutation in homeotic gene result in homeosis, which governs the transformation of structure (one or more) or segment (one or more) into the resemblance of other, but never responsible for the loss of segment and we can say mutation result in ectopic expression of the body part. Gap gene and pair-rule gene in conjugation act as a transcription factor for homeotic gene and act on cis-regulatory elements called initiator enhancer elements. Homeotic gene themselves act as the transcriptional factor and posses MADS domain, in which M denotes MCM, A denotes agamous, D denote deficiently, S denotes SRF).

In Drosophila homeotic gene cluster as HOM-c Box, which divides into two complexes:

Bithorax (BX-C), which themselves contain Ultrabithorax, Abdominal-A and Abdominal B. Fifth to thirteen parasegment identity is controlled by these genes and called a posterior homeotic gene.

Antennapedia (ANT-C), which themselves contain labial (lab), proboscipedia (pb), Deformed (Dfd), Sex comb reduced (Scr) and Antennapedia (Antp). The identity of parasegment present prior to five parasegments get controlled by these genes, called an anterior homeotic gene.

Anterior and posterior homeotic gene represses the expression of each other in their expression region. For eg, Antennapedia represses in ultra biothorax (Ubx) region. If ultra biothorax absent in turn result in one wing now express from T3 just like T2 and homeotic transformation occur. In other cases when Antennapedia mutated or absent, expression of leg occur in place of the antenna. If the mutation in bithorax then T3 show similarity to T2 having a pair of wings along with two abdomens.

HOX is the homologous cluster of the homeotic gene present in the worm to man. There is a precise correlation between the genes expressed along the anterior-posterior axis of the developing embryo and the relative position of each homologue in each of these cluster of the homeotic gene (between HOX and HOM). Only one homeotic gene cluster HOM-C present in primitive insects and the split between Ant-C along with BX-C taken place during insect evolution, thus it shows original cluster form by duplication and the ancestral complex has 8 genes. This complex themselves duplicated in higher metazoans and in duplicated twicely in mammals thus had four clusters (HOX-A to HOX-D). All homeotic gene are the homeodomain-containing transcriptional factor but all homeobox-containing gene are the hot homeotic gene. Homeobox is 60 amino acid or 180 basspair containing conserve box.

Extradenticle (EXD) is a TALE homeodomain protein act as a cofactor for all homeotic gene and both bind to each other through their homeodomain. On double-stranded DNA homeodomain of both protein bind at the opposite side and interact with each other by conserve motif of six amino acid (hexapeptide) located at HOM protein homeodomain N-terminal and EXD homeodomain C-terminal. This interactive model comes in the selective binding model of the homeodomain and reported in fork head (fkh) promoter, between sex comb reduce and extradenticle. One other example of Distal-less gene (Dll) responsible for leg formation in thoracic segments and ultra biothorax along with Aba A control its expression in the abdominal segment, in turn, help the Dll repressor cooperative binding to dsDNA with its cofactor EXD and Homothorax.

According to activity regulation model of the homeodomain, the cofactor is not necessary for the activity of the homeotic gene, this reported in case of Deformed (DFD) and Fushi tarazu when make the association with the powerful transcription activation domain of VP16 protein. HOX gene also possesses cofactor, which includes genes from MEIS and PBC family, thus also show cooperative binding to dsDNA with its cofactor. Mutation in HOXD13 result in synpolydactyly (dominant disorder, which affects digit formation) and mutation in HOXA13 result in hand-foot-genital syndrome.

Mostly HOM gene initially show parasegmental gene expression, sometimes more than one HOM gene also express in a parasegment in combination pattern, in 5-12 parasegment ultra biothorax expression takes place and this is area of gap gene kruppel expression, act as activator for ultra biothorax through categorize the broad central region to which ultra biothorax expression occur. Hunchback and even shipped act as a negative regulator for ultra biothorax. But Fushi tarazu shows positive regulation, by generating pair rule periodicity through strip border sharpening. Hunchback and even shipped make interaction with dMI-2 and RPD-3 respectively, dMI-2 and RPD-3 belongs to histone deacetylase complex in turn help hunchback and even shipped to perform their function. Ultra biothorax promoter contains upstream binding sites for ultra biothorax, even shipped, Fushi tarazu which are found to be closely associated with PREs, TREs, PRE and TRE are those are response element for Polycomb and trithorax gene respectively. Polycomb complex thought to be recruit by even shipped. Largely Fushi tarazu target makes interaction with Fushi tarazu-F1 (orphan nuclear receptor), which recruit histone acetyltransferase.

Autoregulation and interaction through cross-regulation cause additional refinement in the HOM gene after the disappearance of the gap and pair-rule protein. Protein encoded by trithorax (trxG) gene responsible for maintaining the state of transcriptional activation of HOM gene by interacting activator and in turn coactivator which adds acetyl group, thus regulate chromatin remodeling, as well as themselves regulate chromatin remodelling because trithorax possess histone acetyltransferase activity and Polycomb (PcG)  responsible for maintaining the state of transcriptional repression of HOM gene by interacting repressor and in turn corepressor which remove acetyl group thus regulate chromatin remodelling, as well as themselves, regulate chromatin remodelling because polycomb possess histone deacetylase activity and chromodomain, Both genes are two additional classes of genes, responsible for maintaining the transcriptional state of the HOM gene. Chromodomain also present in MOF and MSL-3 (a protein involved in dosage compensation) required to bind roX (structural protein), this RNA target the RNA/protein complex to the male chromosome.

 4.7.        Dorsal-Ventral axis formation:

After fertilization, the nucleus of zygote moves to future dorsal surface. The nucleus transcribes the Gurken mRNA in the dorsal side. Gurken mRNA translate into Gurken protein. This Gurken protein is secreted by oocytes. Gurken protein work as the ligand for torpedo receptor present on the dorsal follicular cell. Torpedo receptor blocks the pipe protein synthesis after binding with Gurken protein. This pipe is not formed at the dorsal surface. Pipe gene expression gets inhibited by torpedo protein at dorsal side, pipe gene plays important role in ventralization of the oocyte surface. Ventralization of the embryo occurs if the mutation or maternal deficiencies take place within either the gurken or the torpedo gene. Through the transplantation experiment, two results come. First, the torpedo mutant eggs were able to produce normal embryos if they transplant into wild type ovary. Second, wild type egg develops mutant, ventralized embryos if they transplanted into the mother's egg chamber, which is torpedo mutant.

4.8.         Ventral axis formation:

Only the ventral follicle cells were able to make the pipe. A protein torpedo does not block pipe synthesis because gurken protein is absent in ventral surface. Slightly later stage of development unknown protein (x) gets modified by pipe protein and both of them get the bind and get secreted from the ventral follicle cells. Pipe activates the Nudel protein. Nudel protein is secreted by ventral embryonic cells and present in the perivitelline fluid.

Three serine proteases which are the product of gastrulation defective (gd), snake (snk), and easter (en) genes and these are secreted within inactive form into the perivitelline fluid, get activated in a cascade manner by nudel.

Activated Nudel activates the Gastrulation-defective protease. The cleaved Gd works as protease and cleaved the snake protein. The cleaved snake works as protease and becomes active. The cleaved snake cleaved the ester protein. Nudel cleaved the inactive Gd. Cleaved Easter protease cause the cleavage of the Spatzle protein. The ventral portion of the embryo is the place where the cleavage of these three proteases is limited.

Spatzle protein is now able to bind to its receptor, which is toll protein (maternal product) present in the oocyte cell membrane throughout and it is the product of the toll gene. Thus, Toll signal only gets receive by the ventral cell. When Spatzle binds to and activates the Toll protein, Toll cause the activation of Pelle, which is a protein kinase. Pelle with the help of Tube, get bring on to the plasma membrane and Tube also get activated here. In the cytoplasm of the syncytial blastoderm, Dorsal-Cactus complex is present. Pelle a protein Kinase cause the phosphorylation of Cactus, as a result, Cactus undergo ubiquitinylation and get degraded. Dorsal this causes the release of ventral side and Dorsal protein moves to nuclei of the ventral part of the embryo. It is a transcription factor, cause the transcription of the gene specifying ventral cell types, as a result, surface get ventralizes.

About 90 minutes after fertilization synthesis of Dorsal protein takes place and present throughout the syncytial blastoderm of Drosophila’s early embryo, as dorsal protein produce instantly form complex with cactus, which is also present in the cytoplasm of the syncytial blastoderm.


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