Date of Award

12-2008

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Legacy Department

Genetics

Advisor

Friez, Michael J

Committee Member

Abbott , Albert G

Committee Member

DuPont , Barbara R

Committee Member

Tomkins , Jeffrey P

Abstract

ABSTRACT
A number of conditions related to X-linked intellectual disabilities (XLID) are in part due to microduplications that are not visible cytogenetically. With the focus on Rho, Ras and Rab genes, a family of genes known to be associated with intellectual disabilities, were screened for dosage aberrations (Leeuwen, F. N. 1997), (Ng, E. L. 2008), (Gissen, P. 2007), (Gurkan, C. 2005). Cohorts of intellectually disabled ID individuals were explored with new technologies. These new technologies include comparative genomic hybridization (CGH), multiplex ligation dependent probe amplification (MLPA) and quantitative PCR (qPCR) (Madrigal, I. 2007), (Hermsen, M. A. 2005), (Morey, J. S. 2006).
The first screening was of two groups of individuals, one group with hypotonia and varying degrees of ID and the other of individuals with nonsyndromic ID and a suspected X-linked etiology. These cohorts were screened using the Mental Retardation on the X chromosome (MRX) kit, which focuses on genes that cause intellectual disability and are located on the X chromosome. The second screening consisted of the two former groups and 5 additional cohorts totaling 1152 patients, using a synthetic probe kit that was designed to target primarily Ras, Rab and Rho X-linked genes that were not covered by the MRX kit. The 5 additional cohorts were individuals that had normal sequencing results for one of the following X-linked genes XNP, L1CAM, UBE3A, FGD1, and STK9.
The MRX screening produced a GDI1 duplication, deletion in FACL4 and an FMR2 missense mutation (c.474C>T). The synthetic MLPA screening found a partial XNP duplication (248kb), a 1p36 duplication/deletion complex rearrangement and a greater than 3Mb 1p36 deletion. It has been concluded from this study that duplications in these genes are rare, appearing in less than 1% in these chosen populations.
Another section of this project is the characterization of a 275kb Xq25 duplication found during routine MLPA testing for MECP2. An Xq25 control peak on the MRC Holland MECP2 MLPA revealed a duplication in a female that presented with a MECP2 phenotype (Chahrour, M. 2007). This duplication spanned four genes (AIF, ELF4, BCORL1 and RAB33A) and of these four, two were over-expressed (AIF and RAB33A). Using qPCR to look for the link that may cause the similar phenotype to Rett syndrome in this patient, 26 Ras, Rab and Rho genes were tested in patients with Rett syndrome, Fragile X syndrome, ID with unknown etiology and the Xq25 patient. A similar pattern of expression was seen in this small cohort with ID. The CREB1 gene, the co-activator of MECP2, part of the transcription factor complex for 21 of the 26 genes screened, plays a role in all of these conditions and may be the linking factor in producing these patterns. The over expression of the AIF gene seemed to play a role in the mis-regulation of many genes, but with uncertainty on how it led to any affect on the phenotype.
In this study duplications that play a role in the causation of ID were found using MLPA technology. As array CGH becomes more refined, with higher coverage and better software, the finding of microduplications that cause ID will increase.

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