生科03級
881618
張庭毓
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生科03級 881618 張庭毓 |
實驗室簡介：
以下是幾篇實驗室的poster:
In early eye development, following the sweep of the morphogenetic furrow (MF), ommatidial pre-clusters differentiate and rotate 90o to establish planar polarity. We found that in the eye discs of hh1 and roDOM mutants in which the MF progression is disrupted, the ommatidial clusters overrotate severely, suggesting that MF progression is required at least at a step to stop the ommatidial rotation at 90o. Several evidences suggested that the ommatidial over-rotation is the result of disruption of MF progression but no the lacking of Hh signaling. The glycoprotein Scabrous (Sca) that is synthesized in the MF and is distributed posteriorly in a string-like structure to ommatidial row 6~8. This pattern is cytochalasin D sensitive, suggesting the involvement of actin. Furthermore, ommatidial clusters posterior to sca mutant clones over rotate in similar degree to those in hh1 mutants. Ectopic expression of Sca posteriorly effectively rescues the over-rotation phenotype in hh1 eye discs. We proposed that concomitant to MF progression, which is regulated by the Hedgehog (Hh) signaling pathway, MF regulates the proper rotation of ommatidial clusters by distributing Sca and perhaps other factors posteriorly.
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During
Drosophila external sensory organ
development, the first step is expression of proneural genes in a small group of
ectodermal cells, known as “proneural clusters” Although each cell in the
proneural cluster is competent to form a neural precursor, these proneural-cluster
cells compete with each other so that a single cell is selected to develop into
a sensory organ precursor (SOP). The remaining cells are prevented from adopting
the SOP neural fate by the process of lateral inhibition that involves cell-cell
interaction mediated by receptor Notch. We show that phyllopod
(phyl), previously identified to be
essential for R7 photoreceptor differentiation, is required for the cell fate
specification of SOP cells. Loss-of-function mutations in phyl result in failure in SOP formation, which leads to missing
bristles in adult flies. Conversely, misexpression of phyl promotes ectopic SOP formation. Therefore, phyl
functions as a genetic switch in specifying the fate of the SOP cells. We found
that the expression of proneural gene is not affected in phyl
mutant, and misexpression of phyl
rescue the bristle-missing defects of proneural mutant, indicating that phyl
functions downstream of proneural genes in specification of SOP cell fate. It
was previously shown that Notch
mutations induce formation of supernumerary SOP cells. We further demonstrate
that phyl acts epistatically to Notch,
and its mRNA level is negatively regulated by Notch signaling. In addition to be regulated by Notch
on the transcriptional level, our recent data suggests that Phyl might
negatively regulate Notch on the protein level. Phyl interacts with Notch in
yeast 2-hybrid assay. Also, overexpression of phyl
shows strong genetic interaction with Notch
in wing vein development. phyl is
specifically expressed in neural cells. To understand how the transcription of phyl
is regulated, promoter analyses are performed. We found that the 3.4 Kb
region upstream of the phyl ORF is sufficient to drive reporter expression similarly as the
endogenous phyl expression. How this region being regulated by different
signaling pathways will be examined.
Protein degradation is essential for many cellular processes. We
have found that sina, a ubiquitin E3
ligase, is involved in es organ development. To further understand it’s
function, an enhancer/suppressor screening is performed by using the deficiency
kit of Drosophila second chromosome.
Also, a yeast two-hybrid analysis will be performed to identify the Sina
interacting proteins. The genes that are identified in both screening will be
studied first.
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multiple
Encodes a Novel Protein in Multiple Dendritic Neurons in the Drosophila
Peripheral Nervous System
The peripheral nervous system of Drosophila embryos is composed of type I and type II neurons. Type I neurons are monodendritic neurons including neurons of external sensory and chordotonal organs, whereas type II neurons are multidendritic (MD) ones. A P-element enhancer trap line, E7-2-36, expresses lacZ in all MD neurons. The annotated gene, CG13143 that locates distally to the P-element insertion site, is expressed in all MD neurons. We named this gene multiple. Several multiple mutants were generated by excision of the P-element. So far, no morphological defect in MD neurons was observed in multiple mutants. Isolation of the full-length cDNA is in progress. multiple-GAL4 transgenic flies will be generated to examine the axonal targeting of MD neurons in the CNS and multiple-GFP transgenic flies will be used to identify the subcellular localization of the Multiple protein. Since MD neurons might respond to touch, thermal and noxious stimuli in the larval stage, behavior assays will be done to test if multiple contributes to the function of MD neurons.
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Eye
Formation Genes eyeless and dachshund are Negatively Regulated by Proneural Genes atonal
and daughterless During Drosophila
Eye Development
In
the Drosophila eye development,
eye master genes eyeless, twin
of eyeless, optix, eye gone, sine
oculis, eyes absent, and
dachshund specify the eye primordium, giving it the potential to form an
eye disc. After eye disc formation, the proneural genes atonal
(ato) and daughterless (da)
that encode basic helix-loop-helix proteins to form heterodimers promote the
differentiation of photoreceptors. In the morphogenetic furrow (MF), where photoreceptor
differentiation initiates, cells express Ato and higher levels of Da. In the
mean time, expression of eyeless, twin of eyeless and optix are
repressed in the MF. sine oculis and eyes absent are highly expressed
both anterior and posterior to, but not in the MF. Expression of dachshund
(dac) is at a high level just ahead of MF and repressed to a basal level
in the MF. Overexpression of eyeless, or combination of eyes absent
and sine ouclis across the MF causes repression of photoreceptor differentiation
in the eye disc and missing of ommatidia in adult compound eye. These results
suggest that negative regulation of eye formation genes in the MF is necessary
for eye development. We observed that the expression of Dac is upregulated in
da or
ato mutant clones in the MF. Coexpression of Da and Ato causes the expression
of Dac being downregulated anterior to the MF. We also observed the same repressive
effect on another eye master gene, eyeless. Right now, we are testing if optix and twin of
eyeless are also repressed by Ato and Da. By in situ hybridization, we showed
that the negative regulation of dac and ey by Ato and Da is at
least partly at the transcriptional level. A fusion protein of Da and the activation
domain of VP16 showed similar repressive effect on Dac, suggesting the Ato and
Da may function as an activator to activate a repressor of the eye formation
genes. We will test whether Drosophila homologues of human TRAP230 and
TRAP240, two components of transcriptional factor complex, are involved in the
regulation of Dac and Ey.
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