Browsing by Subject "cellular and molecular biology"

Sort by: Order: Results:

Now showing items 1-1 of 1
  • Shrestha, Kul (Helsingin yliopisto, 2021)
    Mismatch repair (MMR) proteins are involved in maintaining genome integrity in somatic and germline cells. Defects in MMR leads to various abnormalities, such as tumorigenesis in Lynch syndrome (LS) and infertility. MLH1 (a central MMR protein) along with other MMR proteins are involved in repair of replication-induced errors arising due to inaccurate DNA polymerase. If unrepaired (due to defective MMR), these errors accumulate and give rise to a cellular phenotype known as microsatellite instability (MSI). MSI is a hallmark of tumors associated with LS and results from complete loss of MMR; LS patients have high incidence of colorectal cancer. It is still poorly understood why certain tissues in LS, in particular the gastrointestinal (GI) tract, are more vulnerable to MSI-associated tumors. Also, how heterozygosity of MMR genes impact the germline genomic integrity is not known. We hypothesized that certain tissues, in particular those with high proliferating rates, are vulnerable to MMR associated abnormalities compromising microsatellite stability (prior to complete loss of MMR function) in a tissue-specific manner before tumorigenesis. Our aim was to understand how MMR protein levels contribute to MSI in vivo. In the research presented here, we used Mlh1 heterozygous mice (Mlh1+/- mice) as model of LS to investigate how decreased MLH1 levels contribute to tissue-specific MSI, and whether MSI is detectable prior to loss of MMR function and to neoplastic growth. We tested MSI and measured MLH1 levels in primary cells derived from different organs, focusing on the comparison between highly proliferating small intestine and low proliferating spleen. Further, we tested MSI in sperm cells of Mlh1+/- mice. In addition, we studied the association of loss of heterozygosity (LOH) and Mlh1 promoter methylation to tissue-specific MLH1 expression and MSI. The studies were conducted at 4- and 12-month time point. We discovered that highly proliferating normal tissues (small intestine and sperm) of Mlh1+/- mice display MSI which increases with age, while low proliferating spleen was microsatellite stable. Further, Mlh1+/- small intestine showed sporadic decrease in MLH1 levels which associated with the observed MSI, while Mlh1+/- spleens showed expected MLH1 expression (i.e. approximately 50% of wildtype expression). We observed soma-wide Mlh1 promoter methylation in a subset of Mlh1+/- mice; these mice were the most vulnerable to MLH1 expression level decrease and to MSI in the small intestine, while MLH1 expression in other somatic tissues remained unaffected. In brief, we showed that normal small intestine of Mlh1+/- mice is particularly susceptible to MLH1 depletion giving rise to MSI long before neoplasia. Further, we demonstrated that Mlh1+/- sperm exhibit MSI which associates with germline Mlh1 promoter methylation.