Circadian rhythms are daily rhythms in behavior, metabolism, or other biological activities that persist even in the presence of constant environmental conditions. The oscillators that drive the overt biological rhythms found in virtually all eukaryotes presumably evolved as a result of selection for an internal timekeeping system that would allow anticipation of predictable daily environmental cycles. Abnormalities of circadian processes have been widely observed in patients with major affective disorders. These abnormalities can take the form of a reduction in amplitude of circadian rhythms or of an inappropriate phase relationship of different circadian rhythms to one another. These long-standing observations raise the possibility that circadian abnormalities may be directly related to the pathophysiology or symptomatic expression of major affective disorders. The intriguing connection between major affective disorders and circadian dysfunction cannot be examined in detail until much more is known about the molecular mechanism of the circadian clock in mammals, which at present is entirely obscure. The objective of this research program is to initiate the identification of candidate circadian clock molecules expressed in the mammalian suprachiasmatic nucleus (SCN), the known location of the circadian oscillator. Recently-developed methods for protein-protein interaction cloning allow the isolation of any mammalian clock component that has retained the ability to interact with the Period protein, a circadian clock component from Drosophila that shares with certain mammalian transcription factors a dimerization interface known as the PAS domain. In any complex regulatory loop such as the circadian pacemaker is likely to be, it is probable that the various components interact with a number of positive and negative regulatory factors and effectors---perhaps the best example at present is the regulation of the cell cycle, with its cyclins, cyclin-dependent kinases, phosphatases, kinase inhibitors, and targeted proteolysis. Thus the general goal is to identify any proteins expressed in the SCN that are capable of specifically interacting with various fragments of Per, including (but not limited to) novel PAS dimerization partners. Such proteins could very well represent distant mammalian relatives of circadian regulatory factors in Drosophila. We will use the two-hybrid genetic system to isolate SCN cDNAs encoding proteins that can interact with Period or other PAS proteins. Knowledge of the molecular mechanism of circadian pacemaking in the SCN will provide biochemical and genetic markers for a detailed analysis of the relationship between abnormalities of the circadian system and affective disorders.
