Single-molecule tracking reveals the dynamic turnover of Ipl1 at the kinetochores in .

Aurora kinase B, Ipl1 in , is a master regulator of cell division, required for checkpoint regulation, spindle assembly and disassembly, chromosome segregation, and cytokinesis. Decades of research employed ensemble averaging methods to understand its dynamics and function; however, the dynamic information was lost because of population-based averaging. Here, we use single-molecule imaging and tracking (SMIT) to quantify the recruitment dynamics of Ipl1 at the kinetochores and spindles in live cells.

Autophagy-related protein Atg11 is essential for microtubule-mediated chromosome segregation.

Emerging studies hint at the roles of autophagy-related proteins in various cellular processes. To understand if autophagy-related proteins influence genome stability, we sought to examine a cohort of 35 autophagy mutants in Saccharomyces cerevisiae. We observe cells lacking Atg11 show poor mitotic stability of minichromosomes.

Single-Molecule Tracking dataset for histone H3 (hht1) from live and fixed cells of Schizosaccharomyces pombe.

Single-molecule tracking provides direct real-time measurements of protein dynamics in live cells with high spatiotemporal resolution. In the last decade, this method has been adopted by several labs to understand the dynamics of DNA replication, transcription, telomerase, chromatin remodeling, etc. in a variety of model systems (bacteria, yeast S.

Single-molecule tracking dataset of histone H3 (Hht1) in .

Single-Molecule Tracking (SMT) is a powerful method to quantify protein dynamics in live cells. Recently, we have established a data analysis pipeline for estimating various biophysical parameters (mean squared displacement, diffusion coefficient, bound fraction, residence time, jump distances, jump angles, and track statistics) from the single-molecule time-lapse movies acquired from yeast . We acquired the time-lapse movies using different time intervals (i.

Single-molecule tracking for studying protein dynamics and target-search mechanism in live cells of S. cerevisiae.

Single-molecule tracking (SMT) is a powerful approach to quantify the biophysical parameters of protein dynamics in live cells. Here, we describe a protocol for SMT in live cells of the budding yeast Saccharomyces cerevisiae. We detail how to genetically engineer yeast strains for SMT, how to set up image acquisition parameters, and how different software programs can be used to quantify a variety of biophysical parameters such as diffusion coefficient, residence time, bound fraction, jump angles, and target-search parameters.

In-vivo Single-Molecule Imaging in Yeast: Applications and Challenges.

Single-molecule imaging has gained momentum to quantify the dynamics of biomolecules in live cells, as it provides direct real-time measurements of various cellular activities under their physiological environment. Yeast, a simple and widely used eukaryote, serves as a good model system to quantify single-molecule dynamics of various cellular processes because of its low genomic and cellular complexities, as well as its facile ability to be genetically manipulated. In the past decade, significant developments have been made regarding the intracellular labeling of biomolecules (proteins, mRNA, fatty acids), the microscopy setups to visualize single-molecules and capture their fast dynamics, and the data analysis pipelines to interpret such dynamics.