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Rap1A and rap1B ras-family proteins are prominently expressed in the nucleus of squamous carcinomas: nuclear translocation of GTP-bound active form

Oncogene volume 22, pages 6243–6256 (2003)Cite this article

Abstract

We recently showed that rap1 regulates growth and proliferation in normal keratinocytes, which provoked us to investigate its expression and regulation in malignant cells. Rap1 is variably expressed in whole cell lysates of squamous cell carcinoma (SCC) cell lines. Immunoblot analysis of nuclear and cytosolic fractions and immunohistochemistry revealed that in addition to cytoplasmic expression, SCC cells also exhibit prominent punctate rap1 expression in the nucleus. This unexpected nuclear distribution was confirmed by the evaluation of human oral cancer specimens by immunohistochemistry, which showed both nuclear and cytoplasmic localization. Cytoplasmic rap1 expression was observed mostly in large differentiated cells, whereas nuclear localization was found in morphologically less differentiated cells. Quantitative reverse transcriptase polymerase chain reaction and Northern blot analysis showed that both rap1A and rap1B are expressed in SCC cell lines although rap1B signals are more prominent. Transfection with enhanced GFP-tagged constitutively active and inactive forms of rap1B demonstrated that the active GTP-bound form translocates to the nucleus whereas inactive rap1BGDP is retained in the cytoplasm, much of which is in a perinuclear distribution. Furthermore, growth factors induce nuclear translocation of rap1 in oral cancer cells. This novel discovery that active, GTP-bound rap1 translocates to the nucleus makes it only the second of over 100 small GTP-binding proteins to be identified in the nucleus, and the striking prominence of rap1 expression in the nucleus of SCC cells suggests that activated rap1 plays a role in the malignant process.

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Abbreviations

SMG:

small GTP-binding protein

SCC:

squamous cell carcinoma

RT–PCR:

reverse transcriptase polymerase chain reaction

MAPK:

mitogen-activated protein kinase

GAPDH:

glyceraldehyde-3-phosphate dehydrogenase

NE:

nuclear extract

CE:

cytoplasmic extract

GFP:

green fluorescent protein

EGFP:

enhanced GFP

hnRNP:

heterogenous nuclear ribonucleoprotein

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Acknowledgements

This work was supported in part by a grant from NIH-NIDCR DE00452-01 (NJD) and the University of Michigan's, Head and Neck SPORE grant (1 P50 CA97248).

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Author notes

  1. Raj S Mitra and Zhaocheng Zhang: These authors contributed equally to this manuscript

Authors and Affiliations

  1. Department of Oral Medicine, Pathology and Oncology, University of Michigan School of Dentistry, Ann Arbor, 48109-1078, MI, USA

    Raj S Mitra, Zhaocheng Zhang, Bradley S Henson, Thomas E Carey & Nisha J D'Silva

  2. Departments of Pediatrics and Human Genetics, University of Michigan Medical School, Ann Arbor, 481091, MI, USA

    David M Kurnit

  3. Department of Otolaryngology, Laboratory of Head and Neck Cancer Biology, The University of Michigan Medical School and the Comprehensive Cancer Center, Ann Arbor, 48109-0506, MI, USA

    Thomas E Carey

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  1. Raj S Mitra
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Correspondence to Nisha J D'Silva.

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Mitra, R., Zhang, Z., Henson, B. et al. Rap1A and rap1B ras-family proteins are prominently expressed in the nucleus of squamous carcinomas: nuclear translocation of GTP-bound active form. Oncogene 22, 6243–6256 (2003). https://doi.org/10.1038/sj.onc.1206534

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