UCLA

Among the essential transition metals, iron and zinc are the most studied and much is understood about the effect and regulation of these important cofactors. Yet much still remains unknown about the biochemical mechanisms which are required for cellular homeostasis of these metals. My studies have focused on identifying novel cellular responses to iron toxicity and to zinc deficiency in the budding yeast, Saccharomyces cerevisiae.

The first part of my research focuses on identifying suppressor genes which, when over-expressed, can protect cells against iron toxicity. Suppressor screens were performed using a genomic library in a multicopy vector. The screen identified ORM2 and ORM1 as both were shown to rescue cell growth in high iron conditions. Both Orm1/2 are transmembrane proteins localized at the endoplasmic reticulum (ER). It was previously known that both Orm2 and Orm1 are negative regulators for sphingolipid biosynthesis and that they also play roles in protein quality control. Thus, cells show resistance to iron toxicity when sphingolipid synthesis is inhibited by the over-expression of Orm2/1 or by the treatment with myriocin, a potent inhibitor of sphingolipid synthesis. By measuring sphingolipid LCB levels with and without high iron, high iron was shown to induce sphingolipid biosynthesis and that over-expression of ORM2/1 can reduce sphingolipid levels, which protects cells from iron toxicity. Furthermore, data indicate that iron toxicity is mediated by sphingolipid-activated protein kinases Pkh1 and Ypk1 and by a transcriptional factor Smp1. Contrary to general belief that iron toxicity results from oxidative damage, my study shows that iron toxicity involves activation of sphingolipid signaling.

The second part of my research focuses on elucidation of a novel cellular mechanism which yeast cells have developed in response to zinc deficiency. It was discovered that subunits of RNA polymerase I undergo degradation during zinc starvation. The observed degradation is specific to RNA polymerase I (RNAPI) and its associated proteins. Furthermore, this down-regulation of RNAPI is dependent on vacuolar proteases Pep4 and Prb1. Inactivation of the vacuolar proteases rescues the down-regulations of RNAPI as well as its associated proteins during zinc starvation. RNAPI complex is transported out of the nucleus as a complex and this process is mediated by the exportin Xpo1. Data also suggest that de-ubiquitination of RNAPI is required for its down-regulation during zinc starvation. It is hypothesized that the down-regulation of RNAPI, which is responsible for transcribing ribosomal RNAs and accounts for most transcriptional activities in yeast cells, can conserve energy used for ribosome biogenesis during zinc starvation. Moreover, several subunits of RNAPI are zinc-binding proteins. By degrading RNAPI in the vacuole, yeast cells can conserve and recycle zinc atoms for other important cellular functions during zinc deficiency.