Zinc is an important trace element that is found throughout the human body and functions in structural and catalytic activities. It is the second most abundant transition metal in the body. The highest concentration of Zn+2 is found in the prostate gland. While normal prostate cells have exceptionally high Zn+2 levels it is markedly depleted in prostate cancer. It is not yet clear if Zn+2 reduction is a cause or consequence of prostate cancer but it is important to understand the differences in Zn+2 distribution between noncancerous and cancer cells. The overall goal of this research is to define differences in Zn+2 pools between noncancerous and cancerous prostate cells and correlate these changes with proteins that regulate Zn2+ homeostasis. Because Zn+2 is markedly reduced in prostate cancer we would like elucidate where in the cell Zn+2 is reduced. Cells have the ability to store labile Zn+2 in secretory vesicles and are able to mobilize this metal in response to certain cellular cues, such as stresses or cellular signals. This study aims to reveal the difference in Zn+2 distributions at the subcellular level in noncancerous compared to cancerous cells. The goal is to define the physiological role of Zn+2, as well as determine whether Zn+2 reduction plays a role in the pathology of prostate cancer. This proposal has two specific aims (1) characterize changes in expression levels and/or localization of proteins responsible for controlling Zn2+ homeostasis;(2) elucidate the Zn+2 pools in noncancerous and cancerous cell lines using genetically encoded sensors targeted to different compartment of the cell. These live cell studies will be complemented by x-ray fluorescence microscopy on fixed cells to map total Zn+2 and ICP-MS on subcellular fractions.
Live cell imaging of Zn+2 ions in noncancerous and cancerous prostate cells will greatly enhance our knowledge on how this metal is dysregulated when normal prostate cells become malignant. The tools developed in this work can be targeted to different compartments of the cells to obtain quantitative levels of Zn+2 within individual organelles. Obtaining quantitative Zn+2 levels in different compartments will open new areas of research to investigate what proteins are involved in Zn+2 binding and cellular signals.
|Qin, Yan; Miranda, Jose G; Stoddard, Caitlin I et al. (2013) Direct comparison of a genetically encoded sensor and small molecule indicator: implications for quantification of cytosolic Zn(2+). ACS Chem Biol 8:2366-71|
|Miranda, Jose G; Weaver, Amanda L; Qin, Yan et al. (2012) New alternately colored FRET sensors for simultaneous monitoring of ZnÂ²âº in multiple cellular locations. PLoS One 7:e49371|