Crazy Things Each VAV2 Lover Preferably Should Test Drive

Матеріал з HistoryPedia
Перейти до: навігація, пошук

5��0.6 cells per clone (n=19, p-value Lonafarnib cost on the decreased frequency of clones that still contain an INP). To test if delayed cell cycle progression is sufficient to induce INP differentiation, we over-expressed the Cyclin E/Cdk2-specific inhibitor Dacapo (Dap) in type II lineages. Expression of UAS-dap has previously been shown to delay progression through G1 phase in wing disc cells (Reis and Edgar, 2004). We confirmed that 9D11-GAL4 x UAS-dap delayed INP cell cycle progression and decreased the frequency of INPs in S-phase using the BrdU-EdU pulse-chase assay (data not shown). Similar to tara mis-expression in INPs, ectopic Dap expression caused a decrease in the total number of 9D11-GFP+ cells per brain lobe ( Fig. 8A and B). However, in contrast to tara mis-expression, ectopic Dap did not affect the number of INPs in type II lineages ( Fig. 8A and B). These results demonstrate that delayed progression through G1 phase is not sufficient to induce INP differentiation. To investigate the mechanism by which mis-expression of tara induces INP differentiation, we tested the role Tyrosine Kinase Inhibitor Library cell line of two known SERTAD targets VAV2 (based on mammalian studies): Cdk4 and E2F1/Dp. SERTAD proteins activate or inhibit these targets in a concentration and/or context-dependent manner. To test if inhibition of Cdk4 or E2F1/Dp by Tara might contribute to INP differentiation, we analyzed type II lineages in homozygous mutants of the Cdk43 null allele ( Meyer et al., 2000) or the E2F1i2 hypomorphic allele, which fails to activate transcription of E2F1/Dp target genes ( Royzman et al., 1999). Type II lineages developed normally in Cdk43 mutant larvae ( Fig. 8C and D). In contrast, type II lineages in E2F1i2 mutants had a reduced number of 9D11-GFP+ cells per brain lobe and a reduced number of mature 9D11-GFP+ INPs ( Fig. 8C and D). These type II lineage defects are indistinguishable from the defects observed in tara mis-expressing type II lineages, suggesting that inhibition of E2F1/Dp by tara could cause decreased INP proliferation and increased differentiation. We next tested if elevated expression of Cdk4 or E2F1/Dp could suppress the effects of Tara in INPs. In control larvae, 9D11-GAL4 driven expression of UAS-Cdk4 or combined UAS-E2F1, UAS-Dp, in the absence of tara mis-expression, had no effect on type II lineage development (data not shown). In 9D11-GAL4 x UAS-tara, UAS-Cdk4 larvae, type II lineage development was not significantly different from 9D11-GAL4 x UAS-tara larvae ( Fig.