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Efficiency of nonhomologous DNA end joining varies among somatic tissues, despite similarity in mechanism

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Abstract

Failure to repair DNA double-strand breaks (DSBs) can lead to cell death or cancer. Although nonhomologous end joining (NHEJ) has been studied extensively in mammals, little is known about it in primary tissues. Using oligomeric DNA mimicking endogenous DSBs, NHEJ in cell-free extracts of rat tissues were studied. Results show that efficiency of NHEJ is highest in lungs compared to other somatic tissues. DSBs with compatible and blunt ends joined without modifications, while noncompatible ends joined with minimal alterations in lungs and testes. Thymus exhibited elevated joining, followed by brain and spleen, which could be correlated with NHEJ gene expression. However, NHEJ efficiency was poor in terminally differentiated organs like heart, kidney and liver. Strikingly, NHEJ junctions from these tissues also showed extensive deletions and insertions. Hence, for the first time, we show that despite mode of joining being generally comparable, efficiency of NHEJ varies among primary tissues of mammals.

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Abbreviations

NHEJ:

Nonhomologous end joining

DSB:

Double-strand break

HR:

Homologous recombination

PSL U:

Photo stimulated luminescence unit

BSA:

Bovine serum albumin

DAPI:

4,6-Diamidino-2-phenylindole

PCR:

Polymerase chain reaction

IP:

Immunoprecipitation

RT-PCR:

Reverse transcriptase PCR

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Acknowledgments

We would like to thank Dr. Kumaravel Somasundaram, IISc for providing us with the BRCA1 antibody. We thank Dr. Kavitha C.V. for her help in western blot experiments. We also thank Dr. Binu Tharakan, Abhishek K.V., Mrinal Srivastava, Mridula Nambiar, Nishana M. and other members of SCR laboratory for discussions and help. This work was supported by grant from DAE, India (2008/37/5/BRNS) and IISc start up grant for SCR. We also thank Dr. Raghavan Varadarajan for financial assistance. SS acknowledges Senior Research Fellowship from DBT, India.

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Authors disclose that there is no conflict of interest.

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Authors and Affiliations

  1. Department of Biochemistry, Indian Institute of Science, Bangalore, 560 012, India

    Sheetal Sharma, Bibha Choudhary & Sathees C. Raghavan

  2. Manipal Institute of Regenerative Medicine, Manipal University, Bangalore, 560 071, India

    Bibha Choudhary

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  1. Sheetal Sharma
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Correspondence to Sathees C. Raghavan.

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Sharma, S., Choudhary, B. & Raghavan, S.C. Efficiency of nonhomologous DNA end joining varies among somatic tissues, despite similarity in mechanism. Cell. Mol. Life Sci. 68, 661–676 (2011). https://doi.org/10.1007/s00018-010-0472-x

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