Mammalian cells control lipid homeostasis through the ER membrane bound, basic helix-loop-helix leucine zipper transcription factors called sterol regulatory element binding proteins (SREBPs) (Brown et al., 2000
). SREBPs contain two transmembrane segments and have a membrane topology that places the N- and C-termini in the cytosol separated by a short ~30 amino acid luminal loop. SREBP cleavage activating protein (Scap) binds the C-terminal domain of SREBP and forms a stable complex (Espenshade and Hughes, 2007
). When membranes are replete with sterols, SREBP remains inactive through the ER retention of Scap. Upon depletion of cellular cholesterol, Scap escorts SREBP to the Golgi via COPII transport vesicles where the SREBP N-terminal transcription factor domain is liberated by the sequential action of two Golgi resident proteases. Site-1 protease, a subtilisin-like serine protease, cleaves in the SREBP luminal loop yielding an SREBP N-terminal domain tethered to the membrane through a single transmembrane segment (Brown et al., 2000
). Subsequent intramembrane cleavage by the Site-2 protease, a zinc metalloprotease, releases the soluble SREBP N-terminal transcription factor that travels to the nucleus and upregulates transcription of genes required for lipid synthesis and uptake from the environment. Increased cholesterol supply feeds back to inhibit SREBP-Scap transport to the Golgi, thus maintaining homeostasis.
Fungi contain SREBPs that function as hypoxic transcription factors (Bien and Espenshade, 2010
). The fission yeast Schizosaccharomyces pombe
codes for two SREBP homologs, Sre1 and Sre2 (Hughes et al., 2005
). Sre1 binds to yeast Scap, called Scp1, and the mechanism of sterol regulation is conserved. However, Sre1 functions beyond the control of lipid homeostasis and is a principal hypoxic transcription factor (Todd et al., 2006
). Production of ergosterol, the fission yeast equivalent of cholesterol, is highly oxygen consumptive. Under low oxygen conditions, ergosterol biosynthesis is inhibited and Sre1 precursor is cleaved to produce the active N-terminal transcription factor Sre1N. Thus, ergosterol synthesis serves as an indirect measure of oxygen availability to the cell. In addition, oxygen acts independently to regulate the degradation of Sre1N (Hughes and Espenshade, 2008
). Together these two mechanisms cooperate to regulate production of Sre1N, which is essential for low oxygen growth (Hughes et al., 2005
; Todd et al., 2006
). Sre2 lacks the C-terminal Scp1 binding domain and does not bind to Scp1 (Hughes et al., 2005
). Although the Sre2 transcriptional program has not been determined, Sre2 is not regulated by sterols and is cleaved constitutively. How Sre1 and Sre2 are proteolytically activated is unknown.
The basidiomyceteous fungus Cryptococcus neoformans
is a human opportunistic pathogen that contains a Site-2 protease required for proteolytic activation of C. neoformans
Sre1 (Bien et al., 2009
). However, sequence database searches failed to identify Site-2 protease homologs in fungal ascomycetes encoding SREBP, such as S. pombe
and the pathogen Aspergillus fumigatus
(Bien and Espenshade, 2010
). To identify the Sre1 cleavage machinery, we screened the S. pombe
non-essential haploid deletion collection for genes required to generate Sre1N. We identified four genes defective for SREBP cleavage, dsc1–4
, which define the Golgi Dsc E3 ligase complex. We find that Sre1 and Sre2 cleavage requires the ubiquitin-proteasome system and our data suggest that the Dsc complex may serve a general function in post-ER protein degradation.