A RhoA-FRET Biosensor Mouse for Intravital Imaging in Normal Tissue Homeostasis and Disease Contexts.
Nobis M, Herrmann D, Warren SC, Kadir S, Leung W, Killen M, Magenau A, Stevenson D, Lucas MC, Reischmann N, Vennin C, Conway JRW, Boulghourjian A, Zaratzian A, Law AM, Gallego-Ortega D, Ormandy CJ, Walters SN, Grey ST, Bailey J, Chtanova T, Quinn JMW, Baldock PA, Croucher PI, Schwarz JP, Mrowinska A, Zhang L, Herzog H, Masedunskas A, Hardeman EC, Gunning PW, Del Monte-Nieto G, Harvey RP, Samuel MS, Pajic M, McGhee EJ, Johnsson AE, Sansom OJ, Welch HCE, Morton JP, Strathdee D, Anderson KI, Timpson P
The small GTPase RhoA is involved in a variety of fundamental processes in normal tissue. Spatiotemporal control of RhoA is thought to govern mechanosensing, growth, and motility of cells, while its deregulation is associated with disease development. Here, we describe the generation of a RhoA-fluorescence resonance energy transfer (FRET) biosensor mouse and its utility for monitoring real-time activity of RhoA in a variety of native tissues in vivo. We assess changes in RhoA activity during mechanosensing of osteocytes within the bone and during neutrophil migration. We also demonstrate spatiotemporal order of RhoA activity within crypt cells of the small intestine and during different stages of mammary gestation. Subsequently, we reveal co-option of RhoA activity in both invasive breast and pancreatic cancers, and we assess drug targeting in these disease settings, illustrating the potential for utilizing this mouse to study RhoA activity in vivo in real time.
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Cell reports, 21, 2211-1247, 274-288, 2017
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Truncating PREX2 mutations activate its GEF activity and alter gene expression regulation in NRAS-mutant melanoma.
Lissanu Deribe Y, Shi Y, Rai K, Nezi L, Amin SB, Wu CC, Akdemir KC, Mahdavi M, Peng Q, Chang QE, Hornigold K, Arold ST, Welch HC, Garraway LA, Chin L
PREX2 (phosphatidylinositol-3,4,5-triphosphate-dependent Rac-exchange factor 2) is a PTEN (phosphatase and tensin homolog deleted on chromosome 10) binding protein that is significantly mutated in cutaneous melanoma and pancreatic ductal adenocarcinoma. Here, genetic and biochemical analyses were conducted to elucidate the nature and mechanistic basis of PREX2 mutation in melanoma development. By generating an inducible transgenic mouse model we showed an oncogenic role for a truncating PREX2 mutation (PREX2(E824)*) in vivo in the context of mutant NRAS. Using integrative cross-species gene expression analysis, we identified deregulated cell cycle and cytoskeleton organization as significantly perturbed biological pathways in PREX2 mutant tumors. Mechanistically, truncation of PREX2 activated its Rac1 guanine nucleotide exchange factor activity, abolished binding to PTEN and activated the PI3K (phosphatidyl inositol 3 kinase)/Akt signaling pathway. We further showed that PREX2 truncating mutations or PTEN deletion induces down-regulation of the tumor suppressor and cell cycle regulator CDKN1C (also known as p57(KIP2)). This down-regulation occurs, at least partially, through DNA hypomethylation of a differentially methylated region in chromosome 11 that is a known regulatory region for expression of the CDKN1C gene. Together, these findings identify PREX2 as a mediator of NRAS-mutant melanoma development that acts through the PI3K/PTEN/Akt pathway to regulate gene expression of a cell cycle regulator.
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Proceedings of the National Academy of Sciences of the United States of America, , 1091-6490, , 2016
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