Di-Ras, a Distinct Subgroup of Ras Family GTPases with Unique Biochemical Properties

Abstract

The small GTPase Ras family regulates a variety of cell functions including proliferation and differentiation. Here we have identified novel Ras members, human Di-Ras1 and Di-Ras2, belonging to a distinct branch of the GTPase family. Di-Ras1 and Di-Ras2 specifically expressed in heart and brain share 30 -40% overall identity with other members of Ras family, however, they have the following characteristic substitutions at highly conserved regions among the Ras family. 1) Thr-63 and Ser-65 in Di-Ras are substituted for Ala-59 and Gln-61 positions in Ha-Ras, respectively, that are known to be critical for GTP hydrolysis. 2) Within the effector domains, Di-Ras has Ile at a position corresponding to Asp-33 in Ha-Ras, which is important for its interaction with the downstream effector Raf. As predicted by these substitutions, Di-Ras has only a quite low level of GTPase activity and exists predominantly as a GTPbound form upon its expression in living cells. Moreover, Di-Ras fails to interact with the Ras-binding domain of Raf, resulting in no stimulation of mitogenactivated protein kinase. Interestingly, introduction of Di-Ras into HEK293T cells induces large cellular vacuolation. These findings raise the possibility that Di-Ras might regulate cell morphogenesis in a manner distinct from other members of Ras family.The small GTPase Ras family acts as a molecular switch that regulates a wide range of cell functions including proliferation and differentiation (1-3). Ras is activated in response to a variety of extracellular signals, resulting in stimulation of tyrosine kinases either directly or indirectly. The small GTPase, which cycles between active GTP-bound and inactive GDPbound states, is activated by guanine nucleotide exchange factors that enhance the exchange of bound GDP for GTP and is deactivated by GTPase-activating proteins that increase the intrinsic rate of hydrolysis of bound GTP. The GTP-bound form of Ras associates with several effector molecules, most notably members of the Raf family, the RalGDS family and phosphoinositide 3-kinase. Amino acid residues 32-40 of Ras are important for its interaction with effector molecules and designated as the "effector domain" (4). The biological consequences of their interactions depend greatly on the cell type and on the context of other signaling events.The members of Ras family, at least 13 at present, are characterized by extensive similarities in their effector domains (5). Besides three Ras proteins (Ha-Ras, Ki-Ras, and N-Ras), there are four Rap proteins (Rap1A, Rap1B, Rap2A, and Rap2B), two Ral proteins (RalA and RalB), R-Ras, TC21, R-Ras3/M-Ras, and Rheb in the Ras family. Although many Ras members can interact with the same effector molecules as the three Ras proteins, the physiological roles of most Ras-like GTPases are not fully understood.More recently, some members of the Ras family have begun to be analyzed. For example, Rap1, the closest relative of Ras, has attracted much attention because of the possibility that it functions independently or coor...

Generalized myoclonic epilepsy with photosensitivity in juvenile dogs caused by a defective DIRAS family GTPase 1

“…It is part of the Ras family of small GTPases, which have been linked to many cellular signaling pathways in cell growth and differentiation, synaptic plasticity, learning, and memory (41)(42)(43). DIRAS1 and DIRAS2 form a biochemically and functionally distinct branch of Ras GTPases, which are characterized by a fast guanidine-nucleotide exchange rate (16). The biological function of DIRAS1 in mammals is poorly characterized.…”

“…The expression pattern of the DIRAS1 transcript is poorly characterized and suggested to be limited to the brain and heart (16). We amplified the transcript in 28 canine tissues, including 12 brain regions, the spinal cord, and 15 peripheral tissues, and found abundant expression in all brain regions, whereas the pattern was more limited and variable in extra neural tissues (Fig.…”

RASSF7 negatively regulates pro-apoptotic JNK signaling by inhibiting the activity of phosphorylated-MKK7

“…pmyc-Ha-Ras/G12V was created in a previous study. 35 The point-mutated forms of MKK7 (ST/DE, ST/AA, STS/AAA) and various forms of N-Ras (G12V, DC4 and S17N) were generated by one day PCR as previously described. 36 RNAi.…”

Di-Ras2 Protein Forms a Complex with SmgGDS Protein in Brain Cytosol in Order to Be in a Low Affinity State for Guanine Nucleotides

“…However, Di-Ras proteins fail to interact with the Ras binding domain of Raf or the Ras association domain of RalGDS (6,7). Northern blot analysis has shown that Di-Ras1 mRNA is expressed specifically in brain and heart, whereas Di-Ras2 mRNA is specifically expressed in brain (6).…”

“…However, Di-Ras proteins fail to interact with the Ras binding domain of Raf or the Ras association domain of RalGDS (6,7). Northern blot analysis has shown that Di-Ras1 mRNA is expressed specifically in brain and heart, whereas Di-Ras2 mRNA is specifically expressed in brain (6). We have recently reported that that a Caenorhabditis elegans Di-Ras homolog (drn-1) is expressed specifically in neuronal cells and is involved in the modulation of synaptic activity (8).…”