RIBOSOMAL RNA SEQUENCE ANALYSIS

University Of California, San Francisco, San Francisco CA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Deregulation in cell growth and protein synthesis is an emerging mechanism for cancer formation. A growing list of human syndromes associated with increased cancer susceptibility is characterized by genetic mutations in factors regulating ribosome function. One the best examples is X-linked Dyskeratosis Congenita (X-DC), where the DKC1 gene, encoding for the pseudouridine synthase that modifies rRNA, is found mutated. DKC1 functions within ribonucleoprotein complexes in combination with the box H/ACA small nucleolar RNAs that guide the rRNA site-specific conversion of uridines to pseudouridines, important for the function of the ribosome. However, their specific role in modulating translational control and their impact in human disease is poorly understood. Our laboratory has previously demonstrated, in both a mouse model for X-DC and human X-DC patient cells, that mutations in DKC1 greatly decrease global levels of rRNA pseudouridylation altering translational control of specific mRNAs, which are translated through a distinct mechanism known as IRES (Internal ribosome entry site)-mediated translation. This mode of translation initiation is important to regulate the expression of key mRNAs, including tumor suppressor genes, during distinct cellular events such as cell cycle and cell survival. We hypothesize that impaired rRNA pseudouridylation of specific clusters of pseudouridine residues directly impinge on the ability of the ribosome to engage in IRES-mediated translation initiation of specific mRNAs leading to the pathological features associated with X-DC. The goal of this study is to investigate which pseudouridine residues on the rRNA, catalyzed by DKC1, are required for efficient IRES-mediated translation.