Sterol regulatory element-binding proteins (SREBPs) regulate lipid homeostasis and coordinate sterol metabolism and carotenogenesis in the astaxanthin-producing yeast Phaffia rhodozyma. While Sre1, the SREBP ortholog, and the site-2 protease Stp1 have been identified as essential components of this pathway in P. rhodozyma, additional factors involved in Sre1 processing or regulation remain unknown. In Aspergillus species, a signal peptide peptidase contributes to the activation of the SREBP ortholog, raising the possibility of a similar role in this yeast. In this work, we identified and characterized the P. rhodozyma signal peptide peptidase (SppA) homolog. Sequence analysis, domain prediction, and phylogenetic analyses supported its classification within the SPP family of intramembrane aspartyl proteases. To evaluate its functional role, ΔsppA mutants were constructed in genetic backgrounds with constitutive Sre1 activity, including the cyp61− mutant and a strain expressing the active form of Sre1 (Sre1N). Deletion of SPPA did not alter sensitivity to clotrimazole or cobalt chloride, nor affect pigmentation, indicating that SppA is not required for Sre1 activation in P. rhodozyma. Transcriptomic analyses further showed that expression of SRE1 and of its known target genes remained unchanged upon SPPA deletion. Interestingly, the loss of SppA in the Sre1N background caused marked downregulation of genes associated with protein refolding and unfolded protein binding. In agreement with these transcriptional changes, the Sre1NΔsppA strain displayed increased sensitivity to dithiothreitol. These findings suggest that, although SppA is not involved in Sre1 activation in P. rhodozyma, it may play a role in protein stress-related processes. Future studies will be required to define the molecular mechanisms underlying this role and its integration with protein homeostasis networks.
Baeza et al. (Fri,) studied this question.