Dr. Kim McKim Earns $1.2 Million Grant

The National Institutes of Health has awarded $1,271,000 to Dr. Kim McKim, Principal Investigator at the Waksman Institute, in support of his latest research project "Homolog orientation and segregation in acentrosomal meiosis."

Abstract:

During the first meiotic division, homologous chromosomes linked by chiasmata interact with spindle microtubules and segregate to opposite poles.  Defects in this process lead to aneuploidy in the fertilized egg and usually in death of the developing embryo.  In humans, aneuploidy is a leading cause of spontaneous abortions and infertility in women and causes diseases such as Down, Turner or Klinefelter syndromes.  In many organisms, including mammals and insects, the oocyte meiotic spindle lacks centrosomes.   The absence of centrosomes raises fundamental questions about the assembly and function of meiosis spindles.  In the absence of the microtubule-organizing center found at mitotic spindle poles, the chromosomes generate a signal which stimulates spindle assembly.  It is unclear, however, if the kinetochores play a role in this process.  In cells with centrosomes, the microtubule connections formed between the poles and the kinetochores facilitates orientation of sisters (mitosis) or homologous chromosomes (meiosis I).  In acentrosomal cells, novel mechanisms may be employed to orient the homologs.  We have found that a group of central spindle proteins is critical for formation of a bipolar spindle and orientation of the homologs.  These proteins recruit and organize the interpolar microtubules, a region where antiparallel microtubules overlap, in the center of the spindle.  The microtubules could mediate chromosome behavior by direct lateral interactions or indirectly by interacting with microtubules that make direct contact with the chromosomes. 

In this proposal, we will investigate the mechanisms of homolog orientation in the acentrosomal spindle of Drosophila oocytes.  The CPC is required for meiotic spindle assembly and homolog orientation and is recruited to chromosomes even in the absence of microtubules.  Unlike mitotic cells, CPC proteins are not found at the centromeres.  Instead, the CPC is found in a ring around the chromosomes where it recruits factors which regulate spindle assembly such as Subito.  The ring structure also provides a mechanism for directing spindle bipolarity in the absence of centrosomes.  Since the CPC is recruited to a ring that is not associated with the centromeres, we will determine the factors which recruit the CPC and shape its localization pattern.  To investigate the role of the central spindle in homolog orientation, we will use fluorescently tagged proteins and live imaging to investigate the timing of homolog orientation relative to spindle assembly and establishment of the central spindle.  We will use GFP tagged Cenp-C to mark the centromeres and RFP tagged Aurora B to mark the central spindle.  These experiments will be repeated in mutants defective in spindle assembly or chromosome orientation.  Finally we will carry out a comprehensive analysis of centromere and kinetochore protein function to determine the role of kinetochores in chromosome alignment and segregation.  These experiments will also address the role of the kinetochores in spindle assembly.  Since these genes are essential, we will use sophisticated genetic tools available in Drosophila to generate oocytes lacking these proteins.  This includes newly developed germ line RNAi and germ line clones to test the role of different kinetochore components.