GTP-Binding Protein alpha Subunits

About: GTP-Binding Protein alpha Subunits is a research topic. Over the lifetime, 304 publications have been published within this topic receiving 19915 citations.

Pertussis Toxin ADP-Ribosylation of Apparent GTP-Binding Protein Alpha-Subunits in Spermatozoan Membranes

Abstract: Guanine nucleotide-binding proteins or G proteins have been shown to couple cell-surface receptors to second messenger effector systems such as adenylate cyclase or phosphodiesterase in a variety of cells. G,(N,), Gi(Ni), G,(N,), and transducin, are composed of three subunits, a, j3, and y , and the a subunits are specifically ADP-ribosylated by the bacterial enzymes cholera toxin or pertussis toxin. Resolved by Na.dodecyl.S04 (SDS)-polyacrylamide gel electrophoresis, the (Y subunits have apparent molecular weights (M, ) of 39,000 to 52,000. Kopf and colleagues' and our laboratory2 have shown the presence of pertussis toxin substrates in spermatozoan membranes of M , 39,000 to M , 41 ,000. Here, we show that the structure of these pertussis toxin substrates is similar, but not the same in invertebrate, lower vertebrate, or mammalian spermatozoa. In addition, partial peptide maps of porcine spermatozoan M , 41,000 protein and the a subunit of porcine brain Gi(G,i) are identical. It was shown previously that the chymotryptic map of sea urchin M , 39,000 protein was identical to brain Ciao2.

The N54-αs Mutant Has Decreased Affinity for βγ and Suggests a Mechanism for Coupling Heterotrimeric G Protein Nucleotide Exchange with Subunit Dissociation.

Abstract: Ser54 of Gsα binds guanine nucleotide and Mg2+ as part of a conserved sequence motif in GTP binding proteins. Mutating the homologous residue in small and heterotrimeric G proteins generates dominant-negative proteins, but by protein-specific mechanisms. For αi/o, this results from persistent binding of α to βγ, whereas for small GTP binding proteins and αs this results from persistent binding to guanine nucleotide exchange factor or receptor. This work examined the role of βγ interactions in mediating the properties of the Ser54-like mutants of Gα subunits. Unexpectedly, WT-αs or N54-αs coexpressed with α1B-adrenergic receptor in human embryonic kidney 293 cells decreased receptor stimulation of IP3 production by a cAMP-independent mechanism, but WT-αs was more effective than the mutant. One explanation for this result would be that αs, like Ser47 αi/o, blocks receptor activation by sequestering βγ; implying that N54-αS has reduced affinity for βγ since it was less effective at blocking IP3 production. This possibility was more directly supported by the observation that WT-αs was more effective than the mutant in inhibiting βγ activation of phospholipase Cβ2. Further, in vitro synthesized N54-αs bound biotinylated-βγ with lower apparent affinity than did WT-αs The Cys54 mutation also decreased βγ binding but less effectively than N54-αs Substitution of the conserved Ser in αo with Cys or Asn increased βγ binding, with the Cys mutant being more effective. This suggests that Ser54 of αs is involved in coupling changes in nucleotide binding with altered subunit interactions, and has important implications for how receptors activate G proteins.

GNAQ/11 mutant nonuveal melanoma: same same but different.

Abstract: The work by Livingstone et al. published in this issue of the BJD draws attention to the rarely occurring but prognostically highly relevant GNAQ/11 mutations in nonuveal melanoma. Malignant melanoma harbours certain hotspot mutations in signalling pathways and the occurrence of these mutations differs according to the localization of the primary tumour. In cutaneous melanoma we frequently find mutations in the Ras-Raf-MEK-ERK mitogen-activated protein kinase pathway, most of which are BRAF (~40%) or NRAS (~20%) mutations that occur mutually exclusively, at least in the case of highly active BRAF oncoproteins. These mutations are much less common in acral lentiginous melanoma (ALM), which accounts for 14–15% of BRAF and NRAS mutations. Small molecule drugs inhibiting BRAF-mutated melanoma have been proven effective, leading to a much improved overall survival (OS). Mutations in the proto-oncogene c-KIT can be found in ALM (~12%) and melanoma of the mucous membranes (~16%). Studies investigating the efficacy of KIT inhibitors such as imatinib have shown little to no effect on progression-free survival and OS. Finally, ocular melanoma, which most commonly occurs as uveal melanoma, normally lacks BRAF, NRAS or c-KIT mutations, but frequently (80% of somatic mutations) presents with mutations in GNAQ or GNA11, which are involved in signalling via G-protein-coupled receptors. Additional mutations typically occur in SF3B1, EIF1AX or BAP1 and have been shown to be of prognostic relevance. Metastasized uveal melanoma is known to have a bad prognosis with poor treatment response (e.g. to immunotherapy). In a large analysis of cutaneous melanomas from 318 patients, 52% (n = 166) harboured BRAF somatic mutations, 28% (n = 88) had NRAS somatic mutations and 14% of samples displayed mutations in NF1. Most notably, in the socalled triple wildtype subtype (n = 46), GNAQ (n = 1), GNA11 (n = 2) and KIT (n = 6) mutations were found. Van Raamsdonk et al. sequenced exon 5 of GNAQ and GNA11 in 713 melanocytic neoplasms and exon 4 of GNAQ and GNA11 in 453 of these samples and in all coding exons of GNAQ and GNA11 in 97 uveal melanomas and 45 blue naevi. Of the uveal melanomas analysed, 83% had somatic mutations in GNAQ or GNA11. Of the 273 extraocular melanomas sequenced, only one melanoma located on chronically sundamaged skin showed a mutation in GNAQ, resulting in a frequency of 0 4%. Interestingly, there was no significant difference in outcome on the basis of mutational status (GNAQ, GNA11 or neither mutation) among 81 patients with primary uveal melanoma. Very little is known about GNAQ or GNA11 mutations in nonuveal melanoma. Livingstone et al. retrospectively characterized a cohort of 18 patients with metastasized nonuveal melanomas, all of whom harboured mutations in the GNAQ or GNA11 genes. Molecular characterization of tumour tissues included immunohistochemistry staining, targeted sequencing for known recurrent hotspot mutations and a 130-gene panel to determine tumour mutational burden (TMB). In addition to the GNAQ and GNA11 loci, six SF3B1, three EIF1AX and four BAP1 mutations were detected in this cohort, but had no impact on patient prognosis. The data were correlated with the clinical outcome of these patients. Interestingly, GNAQ/11 mutant nonuveal melanomas behave in a clinically different manner regarding the site of first metastases and show much more ultraviolet-derived mutations compared with uveal melanomas. The TMB was situated right between uveal (low) and cutaneous melanoma (high). However, nonuveal melanomas had a poor response to immune checkpoint inhibitor treatments (such as cytotoxic T-lymphocyte-associated protein 4 inhibitors and programmed cell death protein 1 inhibitors), similar to that seen in uveal melanomas. In conclusion, Livingstone et al. describe a distinct subtype of malignant melanoma that shares the clinical behaviour of the cutaneous and genetic alterations of uveal melanoma. The poor treatment responses render this subtype a candidate for intensive research on therapeutic targets.