GapIII, a new brain‐enriched member of the GTPase‐activating protein family

Abstract

Ras GTPase-activating proteins (GAPs) are negative regulators of ras, which controls proliferation and differentiation in many cells. Ras GAPs have been found in a variety of species from yeast to mammals. We describe here a newly identified mammalian GAP, GapIII, which was obtained by differential screening of a rat oligodendrocyte cDNA library. GapIII putatively encodes a 834 amino acid protein with a predicted molecular weight of 96 kDa, which contains a consensus GAP-related domain (GRD). The protein encoded by this cDNA has high homology with Gap1m, which was recently identified as a putative mammalian homolog of Drosophila Gap1. These proteins contain three structural domains, an N-terminal calcium-dependent phospholipid binding domain, GRD, and a C-terminal PH/Btk domain. Because of the sequence homology and the structural similarities of this protein with Gap1m, we hypothesize that GapIII and Gap1m may be members of a mammalian GAP gene family, separate from p120GAP, neurofibromin (NF1), and IQGAP. To confirm the GapIII protein activity, constructs containing different GapIII-GRD domains were transformed into iral mutant yeast to determine their relative ability to replace IRA1 functionally. Constructs that contained essentially the full-length protein (all three domains), the GRD alone, or the GRD plus PH/Btk domain suppressed heat shock sensitivity of ira1, whereas constructs that contained the GRD with part of the PH/Btk domain had only a weak ability to suppress heat shock sensitivity. These results suggest that the GapIII GRD itself is sufficient to down-regulate ras proteins in yeast. Expression of GapIII mRNA (4.2 kb) was examined by Northern analysis and in situ hybridization. This mRNA was expressed at highest levels in the brain, where its expression increased with development. Lower levels of the mRNA were expressed in the spleen and lung. Among neural cells, GapIII mRNA was expressed in neurons and oligodendrocytes, but not in astrocytes. Interestingly, the expression pattern in brain is reminiscent of type 1 NF1 expression reported by Gutmann et al. (Cell Growth Differ in press, 1995). We propose that in addition to p120GAP and neurofibromin, the GapIII/Gap1m family may be important for modulating ras activity in neurons and oligodendrocytes during normal brain development and in particular in the adult brain.

Up-regulation of ras-GAP genes is reversed by a MEK inhibitor and doxorubicin in v-Ki-ras-transformed NIH/3T3 fibroblasts

“…The 100 kDa Ras-GAP1 family contains two closely related proteins, GAP1 m [15] 3 and GAP1 IP4BP [16] / GAPIII (a mouse homologue of GAP1 IP4BP ) [17]. Though roles of GAP1 are not completely clear, its distinct perinuclear localization [18] leads us to expect that GAP1 m might be involved in the transcriptional process.…”

Genome-wide discovery of Pax7 target genes during development

“…Fold enrichment was determined (expressed as ϩPax7-specific antibody/no antibody) and subjected to unpaired two-tailed t-tests using StatistiXL v1. 3.…”

Reduced expression of neurofibromin in human meningiomas

“…2 and no. 18), their interpretation is complicated by recent identification of other p21 ras-specific GAP proteins in the brain tissue (Maekawa et al, 1994;Weisbach et al, 1994;Baba et al, 1995). At present, it is not known whether these proteins are expressed in meningiomas and whether their GAP activity is susceptible to inhibition by maltoside.…”

“…To eliminate the possibility that other p21 ras-specific GAP molecules (Maekawa et al, 1994;Weisbach et al, 1994;Baba et al, 1995), besides neurofibromin and p120 GAP, contribute to the total and maltoside-sensitive GAP activity of meningioma lysates, neurofibromin and p120 GAP were immunoprecipitated from the soluble fraction of meningiomas, and the GAP activity of immunoprecipitated proteins was determined in p21 ras GTPase assay.…”

“…The NFl gene encodes a 2818 amino acid protein, designated neurofibromin. A 360 amino acid region of neurofibromin shows significant homology to the catalytic domain of the mammalian p21 ras-specific 120-kDa GTPase-activating protein (p120 GAP), yeast equivalents IRAI and IRA2 proteins and recently identified mammalian p21 ras-specific GAPs (Ballester et al, 1990;Maekawa et al, 1994;Weisbach et al, 1994;Baba et al, 1995). This GAP-related domain (GRD) of neurofibromin, as well as the full-length neurofibromin, can negatively regulate p21 ras in vitro by stimulating its weak intrinsic GTPase activity (Xu et al, 1990;Golubic et al, 1992).…”