Combined Micromechanical-Biochemical Study of Mitotic Chromosome Structure

University Of Illinois At Chicago, Chicago IL

Investigators

Abstract

This project is a study of the structure of the mitotic chromosome of animal cells. For this work the investigator will use a unique combination of micromanipulation and biochemical techniques. Biophysical force measurements will be used to assay biochemically-introduced structural changes in mitotic chromosomes. In this way the role of specific types of molecules in defining mitotic chromosome structure can be quantitatively studied. Preliminary results indicate that DNA itself provides the structural integrity of the folded mitotic chromosome. These results challenge the classical protein scaffold model of mitotic chromosome structure, and indicate the value of this approach in ruling out specific models of chromosome structure. The first objective of this project is further study of DNA connectivity in the mitotic chromosome in this way, and to extend these measurements to Xenopus chromosomes. Additional enzyme experiments will study the role of RNA and protein in mitotic chromosome structure. A second objective is to compare the physical properties of Xenopus chromosomes from cells with chromatids reconstituted using Xenopus egg extracts. A third objective is to characterize the interchromosome filaments observed when chromosomes are removed from cells during mitosis, and in particular to quantify their DNA content. The question of whether or not there is a contiguous protein skeleton inside the mitotic chromosome is a basic and open topic in cell biology. Understanding the relation of reconstituted chromosomes to chromosomes in cells is extremely important given the general assumption that the former system is an accurate model of chromosomes. Finally, putting the question of existence of mitotic interchromosome filaments on solid ground will demand new thinking about many assumptions about mitotic chromosome structure. Broader impacts of this research will be the development of new biophysical techniques (nanonewton-scale force measurement, microfluidic enzymatic treatments) for the study of chromosome structure. Along with this, the proposed activities will focus on research education for Ph.D. students, combining physics and biology. There is currently a national shortage of young scientists with this kind of training. This project will therefore uniquely strengthen the scientific personnel and technique pools in this growth area.