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Testing the Role of an Arf GTPase-activating Protein dASAP in Epithelial Cell Polarity in the Drosophila EmbryoShao, Wei 11 January 2011 (has links)
Baz/PAR3 is a key regulator of epithelial cell polarity (ECP). To identify proteins functioning with Baz, I completed a baz genetic interaction screen by localizing 15 GFP-tagged candidates. Then I tested the role of a top candidate, dASAP (Drosophila Arf GTPase-activating protein with SH3 domain, Ankyrin repeat and PH domain), in Drosophila ECP. To determine whether dASAP might interact with polarity players, I defined the localization of dASAP throughout embryogenesis with GFP-tagged proteins and an anti-dASAP antibody. To study how loss of dASAP function affects ECP, I generated a deletion allele by imprecise P-element excision. To evaluate how each of the six domains of dASAP contributes to its localization and functions, I generated constructs deleting each domain. I found associations between dASAP, actin and the apical domain. The six domains may act redundantly to localize dASAP, although interactions between domains may affect the degree of membrane association.
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Testing the Role of an Arf GTPase-activating Protein dASAP in Epithelial Cell Polarity in the Drosophila EmbryoShao, Wei 11 January 2011 (has links)
Baz/PAR3 is a key regulator of epithelial cell polarity (ECP). To identify proteins functioning with Baz, I completed a baz genetic interaction screen by localizing 15 GFP-tagged candidates. Then I tested the role of a top candidate, dASAP (Drosophila Arf GTPase-activating protein with SH3 domain, Ankyrin repeat and PH domain), in Drosophila ECP. To determine whether dASAP might interact with polarity players, I defined the localization of dASAP throughout embryogenesis with GFP-tagged proteins and an anti-dASAP antibody. To study how loss of dASAP function affects ECP, I generated a deletion allele by imprecise P-element excision. To evaluate how each of the six domains of dASAP contributes to its localization and functions, I generated constructs deleting each domain. I found associations between dASAP, actin and the apical domain. The six domains may act redundantly to localize dASAP, although interactions between domains may affect the degree of membrane association.
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The Effect Of Indole Acetic Acid, Abscisic Acid, Gibberellin And Kinetin On The Expression Of Arf1 Gtp Binding Protein Of Pea (pisum Sativum L. Cv. Araka)Ertekin, Ozlem 01 September 2007 (has links) (PDF)
ADP Ribosylation Factor 1 (ARF1) is a universal small GTP binding protein
which has an important role in vesicular trafficking between endoplasmic
reticulum and Golgi. ARF1 is a basic component of Coat Protein I (COPI) vesicles
which have functions in both formation of coatomer complex and recruitment of
cargo proteins. In this study, the expression ARF1 was analyzed in pea (P. sativum
L. cv. Araka) grown at different developmental stages. Because of the differential
hormonal levels at corresponding stages, the effects of hormones on ARF1
expression were also studied.
The results of present research show that ARF1 expression in embryos and 2 days
grown plants after germination is lower when compared to 6 days grown plants. In
order to see the hormonal effect, 3 weeks old plants were supplied with 50µ / M of
each hormone for 3 times on alternate days. Protein extraction, cell fractionation,Western blot was carried out and immunoblot analysis was conducted with
AtARF1 polyclonal antibodies.
It was shown that, in pea shoots, abscisic acid and gibberellin increases the
inactive GDP bound ARF1 by hydrolyzing ARF-GTP through activating ARFGTPase
activating protein (ARF-GAP) or partially inhibiting ARF-Guanine
Nucleotide Exchange Factor (ARF-GEF). In roots, ARF-GDP (cytosolic fraction),
ARF-GTP (microsomal fraction) and total amount of ARF1 (13.000 x g
supernatant fraction) were down regulated by ~11, ~19 and ~11 fold respectively
with the application of gibberellin / and by ~11, ~7 and ~3 fold respectively with
the application of abscisic acid / when compared to control plants. These results
indicate the importance of plant hormones in the regulation of ARF1 in pea.
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