Intriguingly, TOR and RHEB have recently been show to modulate the circadian clock of Drosophila ( Zheng and Sehgal, 2010). However, FKBP proteins have also been implicated

in regulation of nuclear localization and protein stability. For instance, the noncanonical FKBP-like protein (FKBPL) has been implicated in the nuclear import of steroid hormone receptors in complexes with HSP90 proteins (Robson and James, 2012). An interesting possibility is that BDBT is involved in regulating the import of PER/DBT complexes to the nucleus, and that at least some of this regulation is negative, as PER exhibits increased nuclear accumulation in BDBT knockdown flies (Figure 5). This hypothesis is consistent with our structural Enzalutamide supplier work, which uncovered a resemblance between BDBT and the HSP90-binding protein FKBP51. The HSP90-binding site in FKBP51 localizes to its TPR domain and all but one of the residues that account for HSP90 binding are conserved in BDBT in spite of the low sequence homology with BDBT (Figure 7E) (Wu et al., 2004). Since the N-terminal, PPIase-like domain of BDBT binds to DBT in HEK293 cells (Figure 1D), it is possible that BDBT assembles a DBT/PER/HSP90 complex, with DBT bound to the PPIase-like domain, HSP90 to the TPR domain, and PER bound to DBT. FKBPs have also been implicated in

the regulation of the stabilities of proteins with which they form a complex (Kang et al., 2008). A role in enhancement of PER’s phosphorylation-dependent proteolysis is particularly attractive GDC-0449 order for BDBT, as it

would explain the RNAi knockdown phenotype in head extracts (elevated levels of hypophosphorylated PER; Figure 3) and the enhancement of DBT-dependent degradation of PER in S2 cells (Figure 4). The cytosolic BDBT foci in Drosophila photoreceptors accumulate at a time (ZT13-19; Figure 6) when PER transitions from a destabilized cytosolic form to a stabilized nuclear form, and our data supporting Sitaxentan the involvement of BDBT in enhancement of PER proteolysis suggest that BDBT may be a negative regulator of this transition (i.e., BDBT antagonizes PER accumulation and nuclear localization). The BDBT foci are intriguing in light of the finding by Young and coworkers of PER/TIM cytosolic foci, which form prior to accumulation of PER and TIM in S2 cell nuclei ( Meyer et al., 2006). It was proposed that processes in these foci trigger the nuclear accumulation of both PER and TIM. Since the suggestion from our work is that BDBT foci antagonize nuclear accumulation of PER and we do not observe obvious PER foci that colocalize with the BDBT foci ( Figure 6), it is possible that BDBT antagonizes focal accumulation of PER or immediately triggers the degradation of PER in these foci.