Microsatellite binding

Abstract: CD30 is a member of the TNF receptor family. Our interest lies in understanding the control of CD30 expression, particularly as its over-expression provides a diagnostic marker for a subset of non-Hodgkin's lymphomas, particularly anaplastic large cell lymphoma (ALCL), and because anti-CD30 treatment has been shown to be efficacious. We have identified a number of regulatory regions, including an Sp1 element in the minimal promoter, and a downstream promoter element that is required for start site selection. The discovery of both an activating AP1 site and an upstream microsatellite that represses transcriptional activity of CD30 suggests that this region is involved in dysregulation of CD30 expression. We have now identified the major microsatellite binding activity as transcription factor Yin Yang 1 by both one-hybrid cDNA library screening and peptide mass fingerprinting. Due to the strong repressive effect of the microsatellite, we also investigated whether microsatellite instability may induce changes in CD30 expression and hence explain the over-expression of CD30 in ALCL. Laser capture microdissection of ALCL biopsies and CD30 microsatellite typing indicated that the neoplastic cells show a high degree of variation, but this does not correlate with high CD30 expression seen in ALCL.

Abstract: Microsatellite instability (MSI) is important diagnostic and prognostic cancer biomarker. In colorectal, cervical, ovarian and gastric cancers, it can guide prescription of chemotherapy and immunotherapy. In laboratory diagnostics of susceptible tumors, MSI is routinely detected by size of marker PCR products encompassing frequent microsatellite expansion regions. Alternatively, MSI status is screened indirectly by immunohistochemical (IHC) interrogation of microsatellite binding proteins. RNA sequencing (RNAseq) profiling is emerging source of data for wide spectrum of cancer biomarkers. Recently, three RNAseq-based gene signatures were deduced for establishing MSI status in tumor samples. They had 25, 15, and 14 gene products with only one common gene. However, they were developed and tested on the incomplete literature TCGA sampling and never validated experimentally on independent RNAseq samples. In this study, we for the first time systematically validated these three RNAseq MSI signatures on the literature CRC (n=619), endometrial carcinoma (n=533), gastric cancer (n=380), uterine carcinosarcoma (n=55), esophageal cancer (n=83) samples and on the set of experimental CRC RNAseq samples (n=23) for tumors with known MSI status. We found that all three signatures performed well with area under the curve (AUC) range 0.94-1 for the experimental CRCs, and 0.94-1 for the TCGA CRC, esophageal cancer, and uterine carcinosarcoma samples. However, for the TCGA endometrial carcinoma and gastric cancer samples, only two signatures were effective with AUC 0.91-0.97, whereas the third signature showed significantly lower AUC of 0.69-0.88. Software for calculating these MSI signatures using RNAseq data is included.

Abstract: The Rett syndrome protein MeCP2 was described as a methyl-CpG-binding protein, but its exact function remains unknown. Here we show that mouse MeCP2 is a microsatellite binding protein that specifically recognizes hydroxymethylated CA repeats. Depletion of MeCP2 alters chromatin organization of CA repeats and lamina-associated domains and results in nucleosome accumulation on CA repeats and genome-wide transcriptional dysregulation. The structure of MeCP2 in complex with a hydroxymethylated CA repeat reveals a characteristic DNA shape, with considerably modified geometry at the 5-hydroxymethylcytosine, which is recognized specifically by Arg133, a key residue whose mutation causes Rett syndrome. Our work identifies MeCP2 as a microsatellite DNA binding protein that targets the 5hmC-modified CA-rich strand and maintains genome regions nucleosome-free, suggesting a role for MeCP2 dysfunction in Rett syndrome.