EXPRESSION PROFILING REVEALS A POSITIVE REGULATION BY MPER2 ON CIRCADIAN RHYTHM OF CYTOTOXICITY RECEPTORS: LY49C AND NKG2D

Abstract

The mammalian circadian gene, mPer2, an indispensable component of the mammalian circadian clock, not only modulates endogenous circadian rhythms but also plays a crucial role in regulating innate immune function. Previously, we showed that mPer2 plays a crucial role in regulating cytotoxic response. To investigate the molecular mechanism for mPer2-controlled cytotoxic response, in the present study we conducted mRNA expression for 11 genes participating in cytotoxicity regulation in wild-type (WT) and mPer2 knockout (mPer2^{−; ; /  −; ;}) mice bone marrow, that is, Dap-10, Ly49C, Ly49I, Rac1, Mapk1, Map2k1, Nkg2d, Shp-1, Pak1, Pik3ca, and Vav1. The mRNA levels of Ly49C (p < 0.001), Ly49I (p = 0.039), and Nkg2d (p = 0.038) were significantly downregulated in mPer2^{−; ; /  −; ;} mice. Time-dependence of expression profiling was then conducted for four core clock genes (Per1, Bmal1, Clock, Rev-erbα), and six out of these 11 cytotoxic regulation genes (Ly49C, Ly49I, Mapk1, Nkg2d, Shp-1, Pik3ca) in WT and mPer2^{−; ; /  −; ;} entrained in light/dark (LD) or dark/dark (DD) cycles. Consistently, circadian oscillations were observed for Per1, Rev-erbα, Ly49C, and Nkg2d in WT mice under LD and DD cycles. However, these rhythmic expressions were either disrupted or dampened in mPer2^{−; ; /  −; ;} mice. Comparison of gene expression between WT and mPer2^{−; ; /  −; ;} mice showed that mPer2 knockout had systematically downregulated the mRNA expression of two cytotoxicity regulators, Ly49C and Nkg2d. FACS analysis further confirmed that the circadian expression of these genes was not due to the daily difference in cell numbers of NK, NKT, or T cells in bone marrow. Taken together, our results reveal that mPer2 is a critical clock component in modulating circadian rhythms in bone marrow. Furthermore, it implies that Ly49C and Nkg2d are two clock-controlled genes that may play an important role in mediating mPer2-controlled cytotoxic response. (Author correspondence: [email protected])

Keywords:

• Cytotoxicity regulation
• Cytotoxicity receptors
• mPer2
• Ly49C
• Nkg2d

ACKNOWLEDGMENTS

This project is supported by National Natural Science Foundation of China (30770786) and National Natural Science Foundation of China (30628023). We wish to thank Yang Li (Institute of Psychology, CAS) for taking care of the mice and Kai Wang (Aarhus University, DK) for helpful statistical assistance.

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.