Aberrations in fundamental chromosomal processes underlie genetic diseases and cancer in eukaryotic cells. Bacterial pathologies rely on faithful cellular execution of basic chromosomal processes.However, the Kleckner laboratory does not pursue applications of research for commercial purposes. 

• Coordination of DNA replication and cell division in E.coli.
• Recombination-independent pairing of homologous chromosomes.
• Spatial patterning of chromosomal events.
• Chromosome morphogenesis and sister separation in E.coli and mammalian cells.
• Physical basis of chromosome dynamics and spatial patterning along chromosomes.

The Kleckner laboratory is focused on the fundamental processes that govern chromosomal activities. Genetic, biochemical, biophysical, microfludic, molecular and fluorescence and light microscopic imaging methods are used to uncover general principles and specific mechanisms that underlie chromosome function.

E.coli chromosome dynamics. E.coli has long been used as a model organism for elucidation of basic molecular processes. Following this tradition, the Kleckner laboratory uses epifluorescence microscopy of individual living E.coli cells in microfluidic channels at high resolution in time and space, in combination with genetic and molecular analysis, to investigate: (i) the basis of chromosome morphogenesis and sister chromosome separation and (ii) the mechanism(s) by which a cell ensures per-generation coordination between DNA replication and cell division. It is assumed that the principles utilized by E.coli underlie corresponding processes in eukaryotic cells which, in addition, have additional superimposed layers of complexity. Research focuses on the physical principles that underlie these processes, with focus on the possibility that pushing forces play central roles.

Homolog pairing, recombination and spatial patterning of crossovers during meiosis in budding yeast. Sexual reproduction relies on generation of haploid gametes from diploid progenitor cells. This outcome is achieved by meiosis, a modified version of the basic mitotic cell cycle. The central feature of meiosis is a complex program of interactions between homologous maternal and paternal chromosomes (homologs). The Kleckner laboratory studies: (i) the way in which homologous chromosomes "pair", by both recombination-independent and recombination-dependent mechanisms; and (ii) higher order controls of recombination including its specificity for homologs, rather than sister chromosomes, and most especially the mechanism by which crossover recombination events are spatially distributed along chromosomes. Recombination-independent pairing is analyzed in vivo and, in collaboration with the Prentiss laboratory (Department of Physics) in vitro. Crossover patterning occurs in a way that requires communication along the chromosomes. The possibility that this communication could involve redistribution of mechanical stress is of particular interest. An additional problem of interest is how meiotic recombination is biased to occur between homologous chromosomes rather than between sisters. These problems are investigated by genetic approaches combined with molecular analysis, including chromosome capture technology and high resolution two-dimensional gel electrophoresis, and imaging studies of living and fixed chromosomes.

Eukaryotic chromosomes. We are interested in the possibility that pushing forces govern basic eukaryotic chromosome function. One potential manifestation of such effect is the way in which chromosomes progressively acquire organization and structure from "interphase" through metaphase. We study this process by 3D imaging of DNA and chromosomal molecules in combination with genetic and real-time perturbations.

Analysis of objects under conditions of spatial confinement. DNA, chromatin and chromosomes are subject to confinement at multiple levels and length scales. We believe that variations in fiber stiffness, in combination with confinement, generate variations in repulsion/pushing between fiber segments and that these variations underlie many chromosomal processses. To dissect this possibility from one perspective we have created a microfluidic piston in which objects can be subjected to dynamic spatial confinement while remaining accessible to changes in solution milieu, with concomitant visualization of effects by imaging methods of interest. This piston device is being applied to the analysis of diverse chromosome-related objects.

Biological Mechanisms and Pathways

• Chromosome segregation •
• DNA Replication •
• DNA Replication •
• Eukaryotes •
• Genetics •
• Meiosis •
• Molecular biology •
• Physics and mechanics of chromosome dynamics •
• Prokaryotes •
• Recombination
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Infectious Disease

• Escherichia coli (E. coli)
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Research Tools and Instrumentation

• Escherichia coli (E. coli) •
• FACS analysis •
• Fluorescence microscopy •
• Fluorescent In Situ Hybridization •
• Mammalian cells •
• Microfluidics •
• S. cerevisiae
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Therapeutic Discovery Tools and Assays

• Mammalian cells
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