Evolution of Centromeric Protein Coding Genes in the Mus Genus Restricted; Files Only
Jaymes, Gelsey (Spring 2023)
Abstract
Centromeres are chromosomal loci that facilitate error-free chromosome segregation during mitosis and meiosis to avoid aneuploidy and genetic instability, hallmarks of cancers and chromosomal disorders. Paradoxically, while the essential centromere function is strongly conserved, its components (centromeric DNA and associated proteins) are rapidly evolving. Centromeric components provide a unique system to understand evolutionary selection pressure acting on essential genomic components. Our lab has found that genes encoding centromeric proteins are under adaptive Darwinian selection in several Mus species. My study involves investigating the patterns of evolutionary changes in centromeric protein-coding genes in Mus species and how they contribute to the overall evolutionary selection pressure observed in the Mus genus. I found that centromeric proteins evolve differently in different phylogenetic branches of the Mus genus. Furthermore, centromeric proteins show different amounts of localization to centromeres in Mus species suggesting functional evolution in progress. Additionally, toward identifying possible copy number variations, which might be responsible for the rapid evolution of these genes, I have developed a cytological approach to determine the copy number of one of the centromeric protein-coding genes. Understanding the evolution of centromeric protein-coding genes will provide greater insight into centromere function and chromosomal disorders.
Table of Contents
Introduction.................................................................................................................................... 1
Figure 1. Functional centromeres facilitate chromosome segregation during cell division ..................... 3 Results............................................................................................................................................ 3
1. Centromeric protein-coding genes are evolving differently across different phylogenetic branches of
the Mus genus................................................................................................................................. 3
Figure 2. The phylogeny of the Mus genus......................................................................................... 4
Figure 3: Centromeric protein-coding genes are evolving differently across different branches of the Mus genus............................................................................................................................................. 7
2. Centromeric proteins exhibit differences in centromere localization in early and late diverging Mus species........................................................................................................................................... 8
Figure 4. Preparation of cell spreads from mouse liver tissues............................................................. 9
Figure 5. Differences in the localization of centromeric proteins in Mus species. ................................. 10
3. Developing a cytological approach to determine copy number variations for centromeric protein-coding genes............................................................................................................................................. 10
Figure 6. Oligopaint-FISH is a superior cytological visualization method compared to the traditional FISH
bioinformatic approach. ................................................................................................................. 12
Figure 7. Standardization of Oligopaint FISH for HJURP gene in a cell line.......................................... 12
Discussion .................................................................................................................................... 12 Methods........................................................................................................................................ 14
Assembly, alignment, and analysis of centromere protein-coding gene transcripts.............................. 14
Oligopaint HJURP Probe Preparation ............................................................................................... 15
Liver Tissue Cell Spread Production................................................................................................. 17
Oligopaint FISH............................................................................................................................. 18
Immunostaining and Analysis......................................................................................................... 19 References..................................................................................................................................... 20
Supplementary Information ........................................................................................................... 23
Standardization of antibodies against centromeric proteins for immunostaining. ............................... 23
Supplementary Figure 1. Standardization of antibodies against centromeric proteins ......................... 23
Supplementary Table 1. M. spicilegus constructed centromeric protein coding gene transcripts........... 23
Supplementary Table 2. M. macedonicus constructed centromeric protein coding gene transcripts....... 26
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File download under embargo until 19 May 2025 | 2023-04-11 15:28:50 -0400 | File download under embargo until 19 May 2025 |
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