Mechanisms underlying the roles of cell adhesion molecules in the circadian timing system

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The circadian system adjusts internal physiology to the cyclic environment. The light-dark cycle is among the most potent synchronizers to which organisms adjust. In mammals, the circadian system is composed of many clocks coordinating overall physiology and behaviors (eg, activity, feeding, sleep). The suprachiasmatic nucleus (SCN) of the hypothalamus is considered the chief circadian oscillator notably because it directly responds to light. SCN activity/outputs are coordinated by the light-dark cycle and according to the state of cell-cell interactions. Indeed, SCN neurons and glia demonstrate huge variations in their interaction and communication properties. This program proposes that specific cell adhesion molecules (CAMs) involved in neuronal plasticity regulate the circadian system by shaping SCN functioning. We discovered that the CAMs EphA4 and Neuroligin-1 (NLGN1) modulate circadian behaviors. More precisely, we found that mice knockout (KO) for EphA4 and Nlgn1 have a longer endogenous period of wheel-running activity in constant darkness, and that EphA4 KO mice have attenuated circadian responses to light. We now aim at defining whether their roles originate from the SCN, and whether NLGN2 has similar roles. The program will test the hypothesis that different CAMs contribute to SCN functioning via 3 aims: Aim 1 will establish that genetic inactivation of EphA4, NLGN1 and NLGN2 in the mouse SCN disrupt endogenous circadian rhythm, circadian responses to light and circadian flexibility. This will be done by viral delivery of a CRISPR/Cas9 system to independently KO the CAMs in normally developed mice, which will then be submitted to circadian phenotyping using continuous monitoring of wheel-running activity. Aim 2 will determine that genetic rescue of the target CAMs in the SCN of respective KO mice will restore endogenous circadian rhythm, circadian responses to light and circadian flexibility. This will be accomplished by viral delivery of CAMs to the SCN in adult mice KO for the respective CAMs together with circadian phenotyping using wheel-running activity monitoring. Aim 3 will identify the molecular/cellular mechanisms underlying the role of CAMs in SCN functioning using biochemical/molecular biology techniques in mice submitted to SCN CRISPR/Cas9-mediated genetic inactivation or in KO mice submitted or not to genetic rescue. This will include the investigation of different cell types and, in particular, of markers of clock and synaptic function. The program will disentangle the contributions of CAMs to the functioning of the circadian system. It represents a unique area of research and innovates with the methodological approach, the study of the modulatory role of sex, and the investigation of the complex molecular connections between elements of neuronal plasticity and circadian physiology. Importantly, this research assembles an outstanding network of collaborators and will train a diversity of HQP with equity and inclusion.