scholarly journals Aberrant Overexpression of the Rgl2 Ral Small GTPase-specific Guanine Nucleotide Exchange Factor Promotes Pancreatic Cancer Growth through Ral-dependent and Ral-independent Mechanisms

2010 ◽  
Vol 285 (45) ◽  
pp. 34729-34740 ◽  
Author(s):  
Dominico Vigil ◽  
Timothy D. Martin ◽  
Falina Williams ◽  
Jen Jen Yeh ◽  
Sharon L. Campbell ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpase ◽  
Cancer Growth ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

scholarly journals Substrate Trapping Proteomics Reveals Targets of the βTrCP2/FBXW11 Ubiquitin Ligase

2014 ◽  
Vol 35 (1) ◽  
pp. 167-181 ◽  
Author(s):  
Tai Young Kim ◽  
Priscila F. Siesser ◽  
Kent L. Rossman ◽  
Dennis Goldfarb ◽  
Kathryn Mackinnon ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ubiquitin Ligase ◽  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpase ◽  
Mass Spectrometry Analysis ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Defining the full complement of substrates for each ubiquitin ligase remains an important challenge. Improvements in mass spectrometry instrumentation and computation and in protein biochemistry methods have resulted in several new methods for ubiquitin ligase substrate identification. Here we used the parallel adapter capture (PAC) proteomics approach to study βTrCP2/FBXW11, a substrate adaptor for the SKP1–CUL1–F-box (SCF) E3 ubiquitin ligase complex. The processivity of the ubiquitylation reaction necessitates transient physical interactions between FBXW11 and its substrates, thus making biochemical purification of FBXW11-bound substrates difficult. Using the PAC-based approach, we inhibited the proteasome to “trap” ubiquitylated substrates on the SCFFBXW11E3 complex. Comparative mass spectrometry analysis of immunopurified FBXW11 protein complexes before and after proteasome inhibition revealed 21 known and 23 putatively novel substrates. In focused studies, we found that SCFFBXW11bound, polyubiquitylated, and destabilized RAPGEF2, a guanine nucleotide exchange factor that activates the small GTPase RAP1. High RAPGEF2 protein levels promoted cell-cell fusion and, consequently, multinucleation. Surprisingly, this occurred independently of the guanine nucleotide exchange factor (GEF) catalytic activity and of the presence of RAP1. Our data establish new functions for RAPGEF2 that may contribute to aneuploidy in cancer. More broadly, this report supports the continued use of substrate trapping proteomics to comprehensively define targets for E3 ubiquitin ligases. All proteomic data are available via ProteomeXchange with identifier PXD001062.

scholarly journals The Guanine Nucleotide Exchange Factor GBF1 Participates in Rotavirus Replication

2019 ◽  
Vol 93 (19) ◽  
Author(s):  
José L. Martínez ◽  
Francesca Arnoldi ◽  
Elisabeth M. Schraner ◽  
Catherine Eichwald ◽  
Daniela Silva-Ayala ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpase ◽  
Transport Processes ◽  
Replication Cycle ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Virus Surface ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

ABSTRACTCellular and viral factors participate in the replication cycle of rotavirus. We report that the guanine nucleotide exchange factor GBF1, which activates the small GTPase Arf1 to induce COPI transport processes, is required for rotavirus replication since knocking down GBF1 expression by RNA interference or inhibiting its activity by treatment with brefeldin A (BFA) or Golgicide A (GCA) significantly reduces the yield of infectious viral progeny. This reduction in virus yield was related to a block in virus assembly, since in the presence of either BFA or GCA, the assembly of infectious mature triple-layered virions was significantly prevented and only double-layered particles were detected. We report that the catalytic activity of GBF1, but not the activation of Arf1, is essential for the assembly of the outer capsid of rotavirus. We show that both BFA and GCA, as well as interfering with the synthesis of GBF1, alter the electrophoretic mobility of glycoproteins VP7 and NSP4 and block the trimerization of the virus surface protein VP7, a step required for its incorporation into virus particles. Although a posttranslational modification of VP7 (other than glycosylation) could be related to the lack of trimerization, we found that NSP4 might also be involved in this process, since knocking down its expression reduces VP7 trimerization. In support, recombinant VP7 protein overexpressed in transfected cells formed trimers only when cotransfected with NSP4.IMPORTANCERotavirus, a member of the familyReoviridae, is the major cause of severe diarrhea in children and young animals worldwide. Despite significant advances in the characterization of the biology of this virus, the mechanisms involved in morphogenesis of the virus particle are still poorly understood. In this work, we show that the guanine nucleotide exchange factor GBF1, relevant for COPI/Arf1-mediated cellular vesicular transport, participates in the replication cycle of the virus, influencing the correct processing of viral glycoproteins VP7 and NSP4 and the assembly of the virus surface proteins VP7 and VP4.

scholarly journals The guanine nucleotide exchange factor GBF1 participates in rotavirus replication

2019 ◽  
Author(s):  
José L. Martínez ◽  
Francesca Arnoldi ◽  
Elisabeth M. Schraner ◽  
Catherine Eichwald ◽  
Daniela Silva-Ayala ◽  
...  
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Small Gtpase ◽  
Transport Processes ◽  
Replication Cycle ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Virus Surface ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

ABSTRACTCellular and viral factors participate in the replication cycle of rotavirus. We report that the guanine nucleotide exchange factor GBF1, which activates the small GTPase Arf1 to induce COPI transport processes, is required for rotavirus replication since knocking down GBF1 expression by RNA interference, or inhibiting its activity by treatment with Brefeldin A (BFA) or Golgicide A (GCA) significantly reduce the yield of infectious viral progeny. This reduction in virus yield was related to a block in virus assembly since in the presence of either BFA or GCA the assembly of infectious mature triple-layered virions was significantly prevented and only doubled layered-particles were detected. We report that the catalytic activity of GBF1, but not the activation of Arf1, is essential for the assembly of the outer capsid of rotavirus. We show that both BFA and GCA, as well as interfering with the synthesis of GBF1, alter the electrophoretic mobility of glycoproteins VP7 and NSP4 and block the trimerization of the virus surface VP7, a step required for its incorporation into virus particles. Although a post-translational modification of VP7 (other than glycosylation) could be related to the lack of trimerization, we found that NSP4 might also be involved in this process, since knocking-down its expression reduces VP7 trimerizarion. In support, recombinant VP7 protein overexpressed in transfected cells formed trimers only when co-transfected with NSP4.IMPORTANCERotavirus, a member of the family Reoviridae, is the major cause of severe diarrhea in children and young animals worldwide. Despite the significant advances in the characterization of the biology of this virus, the mechanisms involved in morphogenesis of the virus particle are still poorly understood. In this work, we show that the guanine nucleotide exchange factor GBF1, relevant for the COPI/Arf1-mediated cellular vesicular transport, participates in the replication cycle of the virus, influencing the correct processing of viral glycoproteins VP7 and NSP4, and the assembly of the virus surface proteins VP7 and VP4.

scholarly journals Ras-Dependent Regulation of c-Jun Phosphorylation Is Mediated by the Ral Guanine Nucleotide Exchange Factor-Ral Pathway

2000 ◽  
Vol 20 (22) ◽  
pp. 8480-8488 ◽  
Author(s):  
Nancy D. de Ruiter ◽  
Rob M. F. Wolthuis ◽  
Hans van Dam ◽  
Boudewijn M. T. Burgering ◽  
Johannes L. Bos
Keyword(s):  
Guanine Nucleotide Exchange Factor ◽  
Cellular Transformation ◽  
Small Gtpase ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Oncogenic Ras ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Factor C

ABSTRACT The transcription factor c-Jun is critically involved in the regulation of proliferation and differentiation as well as cellular transformation induced by oncogenic Ras. The signal transduction pathways that couple Ras activation to c-Jun phosphorylation are still partially elusive. Here we show that an activated version of the Ras effector Rlf, a guanine nucleotide exchange factor (GEF) of the small GTPase Ral, can induce the phosphorylation of serines 63 and 73 of c-Jun. In addition, we show that growth factor-induced, Ras-mediated phosphorylation of c-Jun is abolished by inhibitory mutants of the RalGEF-Ral pathway. These results suggest that the RalGEF-Ral pathway plays a major role in Ras-dependent c-Jun phosphorylation. Ral-dependent regulation of c-Jun phosphorylation includes JNK, a still elusive JNKK, and possibly Src.

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