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Genes Dev. 2017 Mar 15;31(6):537-552. doi: 10.1101/gad.294413.116.

BRaf signaling principles unveiled by large-scale human mutation analysis with a rapid lentivirus-based gene replacement method.

Author information

1
Department of Pharmacology, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
2
Department of Biological Sciences, Seoul National University, Seoul 08826, Korea.
3
Department of Neuroscience and Cell Biology, Rutgers Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
4
Child Health Institute of New Jersey, Rutgers Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
5
Department of Microbiology, Center for Cell Signaling, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
6
Waisman Center, University of Wisconsin School of Medicine, Madison, Wisconsin 53705, USA.
7
Department of Medical Genetics, University of Wisconsin School of Medicine, Madison, Wisconsin 53705, USA.
8
Department of Neuroscience, Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
9
Undergraduate Class of 2014, Department of Psychology, University of Virginia College of Arts and Sciences, Charlottesville, Virginia 22908, USA.
10
State Key Laboratory of Membrane Biology, School of Life Sciences, Peking University, Beijing 100871, China.
11
Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health, Zhejiang University School of Medicine, Hangzhou 310058, China.
12
Department of Neuroscience, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.

Abstract

Rapid advances in genetics are linking mutations on genes to diseases at an exponential rate, yet characterizing the gene-mutation-cell-behavior relationships essential for precision medicine remains a daunting task. More than 350 mutations on small GTPase BRaf are associated with various tumors, and ∼40 mutations are associated with the neurodevelopmental disorder cardio-facio-cutaneous syndrome (CFC). We developed a fast cost-effective lentivirus-based rapid gene replacement method to interrogate the physiopathology of BRaf and ∼50 disease-linked BRaf mutants, including all CFC-linked mutants. Analysis of simultaneous multiple patch-clamp recordings from 6068 pairs of rat neurons with validation in additional mouse and human neurons and multiple learning tests from 1486 rats identified BRaf as the key missing signaling effector in the common synaptic NMDA-R-CaMKII-SynGap-Ras-BRaf-MEK-ERK transduction cascade. Moreover, the analysis creates the original big data unveiling three general features of BRaf signaling. This study establishes the first efficient procedure that permits large-scale functional analysis of human disease-linked mutations essential for precision medicine.

KEYWORDS:

BRaf; CFC syndrome; cancer; neurodevelopmental disorder; synaptic transmission

PMID:
28404629
PMCID:
PMC5393050
DOI:
10.1101/gad.294413.116
[Indexed for MEDLINE]
Free PMC Article

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