Supporting Data for: Differential expression of VA opsin transcript variants in tissues linked to photoperiodic time measurement in Svalbard rock ptarmigan

Abstract: Photoperiodic time measurement is the process through which annual changes in daylength are integrated to synchronize seasonal physiological and behavioral responses. Unlike mammals, which detect light exclusively through the eyes, birds possess multiple photoreceptive sites including the eyes, pineal gland, and deep brain photoreceptors (DBPs) located within the hypothalamus. The cellular and molecular identity of the DBPs mediating avian photoperiodic time measurement remains a matter of debate, however. Vertebrate ancient opsin (VA opn) is considered a strong DBP candidate, based on its anatomical localization and spectral response characteristics. Here, we used three different RNA profiling methods to evaluate VA opn expression across photosensitive structures in the Svalbard rock ptarmigan, a highly photoperiodic galliform species adapted to the extreme light conditions of the High Arctic. In the pineal gland, over 60% of transcripts encoded a full-length VA opn, containing all seven transmembrane domains considered necessary for photoreceptor function. In vitro analysis demonstrated that the full-length VA opn has peak spectral sensitivity at approximately 500 nm, consistent with published work in other galliform species. By contrast, in the hypothalamus and retina, the majority of VA opn transcripts are predicted to encode non-functional proteins lacking at least 2 out of 7 transmembrane domains. In the hypothalamus, single-nuclei RNA-sequencing analysis showed that VA opn transcript variants are predominantly expressed in cells of the oligodendrocyte lineage. These data support a model in which, in the Svalbard rock ptarmigan, VA opn contributes to the photosensitivity of the pineal gland, but not to hypothalamic DBP function.