cysteine-rich with EGF-like Domains (creld) genes encode evolutionary conserved proteins. Two paralogues are identified in mammals: Creld1 was shown to act as a regulator of the calcineurin/NFATc signaling pathway, whereas creld2 is involved in ER stress response.
The Drosophila genome encodes only one creld gene yielding two splice variant proteins (46% similarty to mammalian CRELD1 and 49% similarty to CRELD2). In order to elucidate the function of Drosophila Creld proteins, we generated a Drosophila null mutant for creld and analyzed the ensuing phenotypes.
As the mammalian orthologues, Creld is localized to the ER membrane. However, we could not confirm an upregulation of gene expression under ER stress of either Creld isoforms, neither in wild type flies nor Schneider cells. Addtionally, creld mutants are able to survive under ER stress (induced by feeding tunicamycin and thapsigargin), indicating that the knock out of Creld is not provoking ER stress sensitivity.
Creld mutant flies suffer from extensive neurodegeneration, accompagnied by severe locomotor and flight defects and reduced viability, but fertile. q-RT-PCR analysis of adult flies could identify a decreased relative mitochondrial DNA content in creld mutants compared to wild type flies, suggesting problems with mitochondrial biogenesis or quality control. Additonally, a decreased number of mitochondria was confirmed in larval tissue using immunofluorescent techniques. The observed phenotypes could significantly be rescued by complementing the food with mitochondrial activity inducing drugs. We performed climbing assays with mutant flies fed with these drugs and were able to see significantly increased climbing capabilites of those flies compared to mutants fed with normal yeast food without any addtives. But no indication of oxidative stress or increased ROS production was observable in creld mutant flies.
We hypothesize that the observed neurodegeneration is at least partly induced by energy deprivation that goes along with reduced mitochondrial mass, and which was evident by increased AMPK phosphorylation.
We conclude from our data that Creld might be involved in mitochondrial biogenesis and/or quality control. Due to its localization to the ER membrane, it might pose a link between the ER compartment and mitochondria, which are closely connectet in order to regulate Ca2+ signaling, regulating intracellular trafficking and controlling mitochondrial biogenesis. Future research will adress the function of Creld in this context.