The Rap1 GTPase functions as a regulator of morphogenesis invivo

Abstract

The Ras-related Rap GTPases are highly conserved across diverse species but their normal biological function is not well understood. Initial studies in mammalian cells suggested a role for Rap as a Ras antagonist. More recent experiments indicate functions in calcium-and cAMP-mediated signaling and it has been proposed that protein kinase A-mediated phosphorylation activates Rap in vivo. We show that Ras1-mediated signaling pathways in Drosophila are not influenced by Rap1 levels, suggesting that Ras1 and Rap1 function via distinct pathways. Moreover, a mutation that abolishes the putative cAMP-dependent kinase phosphorylation site of Drosophila Rap1 can still rescue the Rap1 mutant phenotype. Our experiments show that Rap1 is not needed for cell proliferation and cell-fate specification but demonstrate a critical function for Rap1 in regulating normal morphogenesis in the eye disk, the ovary and the embryo. Rap1 mutations also disrupt cell migrations and cause abnormalities in cell shape. These findings indicate a role for Rap proteins as regulators of morphogenesis in vivo.

Quantitative determination of Rap 1 activation in cyclic nucleotide-treated HL-60 leukemic cells: lack of Rap 1 activation in variant cells

“…For example, Rap 1 mRNA increases during phorbol esterinduced di erentiation of HL-60 cells (Adachi et al, 1992;Yamashiro et al, 1993) and HL-60 cells express two di erent forms of RapGAP, p130 RapGAP and p85/95 RapGAP; p130 RapGAP decreases during phorbol ester-induced di erentiation of HL-60 cells and p85/95 RapGAP decreases during dimethyl sulfoxide-induced di erentiation of HL-60 cells (Kurachi et al, 1997;Rubinfeld et al, 1991). Rap 1 has also been shown to regulate di erentiation in Drosophila photoreceptor cells (Hariharan et al, 1991;Asha et al, 1999) and Rap 1B is required for cAMP-induced di erentiation of PC12 cells (Vossler et al, 1997).…”

The Crk signaling pathway contributes to the bombesin-induced activation of the small GTPase Rap1 in Swiss 3T3 cells

“…In contrast, Rap1 itself was shown to induce DNA synthesis, cellular transformation and tumor formation in nude mice (Altschuler and Ribeiro-Neto, 1998;Yoshida et al, 1992). In Drosophila, deletion of both rap1 alleles is lethal, and a reduced level of Rap1 results in impaired development of the eyes and the ovaries, but also in developmental defects of the embryos, independently of Ras signaling pathways (Asha et al, 1999;Hariharan et al, 1991).…”

Novel Rap1 dominant-negative mutants interfere selectively with C3G and Epac

“…They were lysed with detergents and a portion of the crude lysate was removed to determine the total level of expressed Rap1; the lysate was then centrifuged and processed as in Figure 4 to measure the levels of Rap1-GTP and soluble Rap1. (Hariharan et al, 1991;Asha et al, 1999). Moreover, experiments with mutant somatic cell clones have shown that D-Rap1 is required for viability in a cell-autonomous manner (Asha et al, 1999).…”

“…(Hariharan et al, 1991;Asha et al, 1999). Moreover, experiments with mutant somatic cell clones have shown that D-Rap1 is required for viability in a cell-autonomous manner (Asha et al, 1999). We generated transgenic flies containing the DRap1[S17A] transgene under the control of a UAS sequence and used the Gal4-UAS transcription system to activate the expression of this transgene (Brand and Perrimon, 1993).…”

“…A minimum of 10 independent insertions were generated for each transgene. UAS-DRap1[V12] and UAS-D-Rap1 (Asha et al, 1999) were generously provided by I Hariharan, and UAS-C3G MT3 (Ishimaru et al, 1999) by U Gaul. EP(2)0388 (a strain containing pUAS-D-PDZGEF) was obtained from the Szeged stock centre and UAS-Ras1 (BL-5788) (Karim and Rubin, 1998) from the Bloomington stock centre.…”