Doctoral Dissertations

Date of Award

8-2011

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biochemistry and Cellular and Molecular Biology

Major Professor

Bruce D. McKee

Committee Members

Elias Fernandez, Hong Guo, Jae Park, Mariano Labrador, Todd Reynolds

Abstract

Drosophila male is an example of achiasmatic meiosis which lacks crossingover and chiasmata during meiosis. Previous studies showed that homologous pairing of both euchromatin and centromeres is lost during middle prophase I, however, homologs are still connected as they form bivalents. The X-Y pair utilizes a specific repeated sequence within the heterochromatic ribosomal DNA blocks as a pairing site. No pairing sites have yet been identified for the autosomes. To search for such sites, we utilized probes specifically targeting heterochromatin regions to assay pairing sequences and behavior in meiosis by fluorescence in situ hybridization (FISH). We found that the fourth homologs pair at the heterochromatic region 61 and associate with the X chromosome throughout prophase I. The pairing of the fourth homologs is disrupted in the homolog conjunction complex mutants. Conversely, six tested heterochromatic regions of the major autosomes (second and third chromosomes) have proved to be largely unpaired after early prophase I. This suggests that pairing mechanism of the major autosomes may differ from the sex and fourth chromosomes; stable connections between major autosomal homologs might occur at different sites along chromosomes in different cells by analogy to chiasmata. Moreover, FISH analysis also revealed two distinct patterns of sister chromatid cohesion in heterochromatin: regions with stable cohesion and regions lacking cohesion, suggesting that sister chromatid cohesion is incomplete within heterochromatin but with preferential sites in male meiosis.
Modifier of Mdg4 in Meiosis (MNM) and Stromalin in Meiosis (SNM) are components of homolog conjunction complex and essential for homolog pairing and segregation in male meiosis. Using yeast two-hybrid assay and co-immunoprecipitation, we showed that the MNM and SNM interact with each other. Specifically, the BTB domain of MNM is responsible for the interaction with SNM, whereas FLYWCH domain of MNM is crucial for this interaction but does not directly interact with SNM. Additionally, point mutation analysis revealed that L9K replacement of the BTB domain weakened the MNM-SNM interaction and caused high frequencies of chromosome nondisjunction.
In conclusion, these results provide a biochemical basis for the mechanism of homolog pairing and support the role of homolog conjunction complex in male meiosis.

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