Investigation of cell cycle network of yeast using lab-on-a-chip platform

Abstract

Genome instability has long been implicated as a salient causal factor in aging and age related diseases such as cancer and neurodegeneration. However, the molecular mechanisms associated with genome instability remain unclear. Recent studies suggest that growth signaling in the organism Saccharomyces cerevisiae and in higher eukaryotes might affect oxidative stress and aging/age-related diseases by activating DNA replication stress that causes DNA damage. Within the scope of this thesis, to investigate the role of Sld7p in yeast cells, a network biology approach, combining the interactome and transcriptome data of budding yeast, was applied. The protein-protein interaction networks of cell cycle and chronological aging processes in budding yeast were reconstructed. The ultimate aim was to determine the putative molecular function of the Sld7p by using microfluidic technology, where the behavior of the wild-type and sld7Δ mutant cells were followed on a Lab-on-a-Chip (LoC) platform under different conditions and consequently to support the computational studies by experimental findings. The wild type and mutant yeast cells were grown in glucose rich YNB or YPD media with or without an alkylating agent. The brightfield microscopy images of the trapped cells were captured by Nikon Eclipse-T inverted microscope at regular time intervals and then processed via ImageJ software. The results of cell count, area and perimeter values were analyzed and used to calculate the cell sizes (mother and bud). The durations of G1 and S/G2/M phases of wild type and sld7Δ mutant cells were determined under control and environmental stress conditions. Our multi-omics approach indicated a dual role for Sld7: it participates in “macromolecular complex binding” in cell cycle and has “oxidoreductase activity” in chronological aging in budding yeast. These potential functions of Sld7 in yeast can offer new insights on the role of its putative homolog MTBP in humans.