Mutations in RABL3 alter KRAS prenylation and are associated with hereditary pancreatic cancer

Abstract

Pancreatic ductal adenocarcinoma is an aggressive cancer with limited treatment options1. Approximately 10% of cases exhibit familial predisposition, but causative genes are not known in most families2. We perform whole-genome sequence analysis in a family with multiple cases of pancreatic ductal adenocarcinoma and identify a germline truncating mutation in the member of the RAS oncogene family-like 3 (RABL3) gene. Heterozygous rabl3 mutant zebrafish show increased susceptibility to cancer formation. Transcriptomic and mass spectrometry approaches implicate RABL3 in RAS pathway regulation and identify an interaction with RAP1GDS1 (SmgGDS), a chaperone regulating prenylation of RAS GTPases3. Indeed, the truncated mutant RABL3 protein accelerates KRAS prenylation and requires RAS proteins to promote cell proliferation. Finally, evidence in patient cohorts with developmental disorders implicates germline RABL3 mutations in RASopathy syndromes. Our studies identify RABL3 mutations as a target for genetic testing in cancer families and uncover a mechanism for dysregulated RAS activity in development and cancer.

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Fig. 1: Whole-genome sequencing in a family cluster of pancreatic cancer identifies a germline nonsense mutation in RABL3.
Fig. 2: RABL3 mutation promotes cell proliferation in vitro and cancer in zebrafish.
Fig. 3: RABL3 mutation dysregulates KRAS activity.
Fig. 4: Homozygous rabl3-TR mutants resemble human RASopathy syndromes.

Data availability

RNA-seq data are available through GEO under accession GSE129081. Interacting proteomic data are available through Peptide Atlas under accession PASS01355. Additional data generated in this study are available within the paper and in the supplementary information.

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Acknowledgements

This work was supported by the NIH grant nos. K08 DK105326 (to S.N.); R01 DK090311, R01 DK095721 and R24 OD017870 (to W.G.); R01 GM095567, R01 CA157490, R01 CA188048, P01 CA117969 and R35 CA232124 (to A.C.K.); R01 CA188871 (to C.W.) and R01 GM040602 (to C.A.F.); as well as grants from the National Pancreas Foundation (to S.N.), the Harvard Digestive Diseases Center (grant no. P30 DK034854 to S.N. and W.G.), the Ken and Louise Goldberg Award (to S.N.), an ACS Research Scholar Grant (RSG-13-298-01-TBG to A.C.K.), the Lustgarten Foundation and SU2C (to A.C.K) and the Anna Fuller Fund and the Claudia Adams Barr Program for Innovative Cancer Research (to W.G.). S.N. is a recipient of the Burroughs Wellcome Fund Career Award for Medical Scientists. W.G. is a Pew Scholar in the Biomedical Sciences.

Author information

S.N. and W.G. conceived and designed the overall project. S.S. and C.I.U. assisted with selecting the family, gathering the clinical histories and collecting DNA samples under human subject IRB-approved protocols. S.N., W.G. and I.L. designed the WGS analysis. I.L. performed the WGS analysis and candidate variant filtering. S.N., J.W., A.J.K., J.E.H., A.G.C. and J.H. designed and generated the zebrafish rabl3 mutant lines and performed the cancer studies. J.R.H. and S.N. performed zebrafish histology preparation and analysis. J.D.M. performed and analyzed the AP–MS experiments and CompPASS suite protein interactomics. S.N., W.G. and C.W. conceived and designed the in vitro immunoprecipitation, prenylation assays and HEK293T cell proliferation assays, and P.G., A.B., E.L. and B.U. performed these experiments. S.N. and O.M. designed and performed RASless MEF experiments. J.W.P. performed protein structural modeling. B.C.J. and C.A.F. designed and performed purification of recombinant protein. J.A.P., S.G. and J.D.M. assisted with mass spectrometry analysis. Y.H. assisted with RNA-seq data analysis. M.B.G. performed the zebrafish µCT and bone histomorphometric analysis. O.M., X.W. and J.D.M. provided assistance with tissue culture experiments. C.A.C. and J.A.R. provided analysis of clinical exome sequencing data. C.A.C. and I.L. provided analysis of variants in the Exome Aggregation Consortium. J.W.H., G.G., S.R.S., K.C. and A.C.K. provided overall input. S.N. and W.G. wrote the manuscript. All authors reviewed and edited the manuscript.

Correspondence to Wolfram Goessling.

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Competing interests

A.C.K. has financial interests in Vescor Therapeutics, LLC. A.C.K. is an inventor on patents pertaining to Kras-regulated metabolic pathways, redox control pathways in pancreatic cancer, targeting GOT1 as a therapeutic approach and the autophagic control of iron metabolism. A.C.K. is on the SAB of Cornerstone/Rafael Pharmaceuticals. G.G. receives research funds from IBM and Pharmacyclics. W.G. receives patent royalties from FATE Therapeutics and is on the SAB of Camp4 Therapeutics.

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Supplementary information

Supplementary Information

Supplementary Figs. 1–19 and Supplementary Tables 3 and 5

Reporting Summary

Supplementary Table 1

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Supplementary Table 4

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