Hemanth Nag Noothalapati Venkata
Shimane University, Japan
Title: Stable isotope probing coupled Raman microscopy: An efficient way to study single cell biochemistry
Biography
Biography: Hemanth Nag Noothalapati Venkata
Abstract
Lipid droplets have been hypothesized to be intimately associated with intracellular proteins. However, there is little direct evidence for both spatiotemporal and functional relations between lipid droplets and proteins provided by molecular -level studies on intact cells. To elucidate the interplay between them at the single cell level, Raman microscopy was coupled with a very powerful strategy, namely, stable isotope labeling. Here, I present in vivo time lapse Raman imaging, coupled with stableisotope (13C) labeling, of single living Schizosaccharomyces pombe cells. Our results show that the proteins newly synthesized from incorporated 13C-substrate are localized specifically to lipid droplets as the lipid concentration within the cell increases. Lipids, which help to store energy in a compact form, have variety of roles in biological systems and their metabolism is centralto life. Here, we show that combination of stable isotope probing (SIP), Raman micro-spectroscopy and multivariate curve resolution analysis can serve as a valuable approach in metabolomics research. We studied ergosterol biosynthesis in single living fission yeast cells, grown in mixtures of normal (12C) and 13C-glucose as the sole carbon source. By carefully looking into the biosynthetic pathways and by comparing the observed peak positions with calculation results on isotope-substituted ergosterol, it is possible to understand how 13C is incorporated in the conjugated C=C moiety of the molecule. The multivariate spectral data analysis revealed intrinsic spectra and their relative abundances of all isotopomers.