Parag-Sharma, K. | Search bioRxiv

bioRxiv2020

Optogenetic activation of heterotrimeric Gi proteins by LOV2GIVe - a rationally engineered modular protein

Heterotrimeric G-proteins are signal transducers that mediate the action of many natural extracellular stimuli as well as of many therapeutic agents. Non-invasive approaches to manipulate the activity of G-proteins with high precision are crucial to understand their regulation in space and time. Here, we engineered LOV2GIVe, a modular protein that allows the activation of Gi proteins with blue light. This optogenetic construct relies on a versatile design that differs from tools previously developed for similar purposes, i.e. metazoan opsins, which are light-activated GPCRs. To make LOV2GIVe, we fused a peptide derived from a non-GPCR protein that activates Gi (but not Gs, Gq, or G12) to a small plant protein domain, such that light uncages the G-protein activating module. Targeting LOV2GIVe to cell membranes allowed for light-dependent activation of Gi proteins in different experimental systems. In summary, LOV2GIVe expands the armamentarium and versatility of tools available to manipulate heterotrimeric G-protein activity. GRAPHICAL SUMMARY O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/253781v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@1870610org.highwire.dtl.DTLVardef@19f2b2dorg.highwire.dtl.DTLVardef@cb9e57org.highwire.dtl.DTLVardef@88924_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2020

CRTC1-MAML2 Establishes a PGC1α-IGF1 Circuit that Confers Vulnerability to PPARγ Inhibition

Mucoepidermoid carcinoma (MEC) is a life-threatening salivary gland cancer that is driven primarily by the transcriptional co-activator fusion CRTC1-MAML2. The mechanisms by which the chimeric CRTC1-MAML2 oncoprotein rewires gene expression programs that promote tumorigenesis remain poorly understood. Here, we show that CRTC1-MAML2 induces transcriptional activation of the non-canonical PGC-1 splice variant PGC-14, which regulates PPAR{gamma}-dependent IGF-1 expression. This mitogenic transcriptional circuitry is consistent across cell lines and primary tumors. CRTC1-MAML2 positive tumors are dominated by IGF-1 pathway activation and small molecule drug screens reveal that tumor cells harboring the fusion gene are selectively sensitive to IGF-1R inhibition. Furthermore, this dependence on autocrine regulation of IGF-1 transcription renders MEC cells susceptible to PPAR{gamma}inhibition with inverse agonists. These results yield insights into the aberrant co-regulatory functions of CRTC1-MAML2 and identify a specific vulnerability that can be exploited for precision therapy.

cancer biology↗