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HnRNP L stimulates splicing of the eNOS gene by binding to variable-length CA repeats

Abstract

In the human genome, dinucleotide repeats are common sequence elements of unknown functional significance. Here we demonstrate that CA repeats in intron 13 of the human endothelial nitric oxide synthase (eNOS) gene function as an unusual intronic splicing enhancer, whose activity depends on the CA repeat number. We identify the 65 kDa heterogenous nuclear ribonucleoprotein (hnRNP) L as the major factor that binds specifically and in a length-dependent manner to the CA-repeat enhancer. In addition, we show that hnRNP L functions as a specific activator of eNOS splicing, providing the first evidence for a role of hnRNP L in the regulation of mRNA splicing. We hypothesize that hnRNP L may be involved in the regulation of many other genes containing CA repeats or A/C-rich enhancers.

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Figure 1: CA repeats function in vitro as a length-dependent intronic splicing enhancer.
Figure 2: Splicing enhancer activity of CA repeats in vivo.
Figure 3: Identification of hnRNP L as major CA-repeat–binding factor by UV crosslinking and RNA affinity purification.
Figure 4: hnRNP L is specifically required for eNOS pre-mRNA splicing.

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Acknowledgements

We thank R. Robinson and J. Neveu for expertise in mass-spectrometry and peptide sequencing, G. Dreyfuss for monoclonal hnRNP L antibody 4D11, J. Kim and M. Swanson for constructs, and E. Stickeler for polyclonal YB-1 antiserum. We also thank G. Reither for unpublished data, S. Schreiner for technical assistance and Z. Palfi for comments on the manuscript. We are grateful to P. Kloetzel (Charité, Berlin) for providing laboratory space during the initial phase of this work. This work was supported by the Deutsche Forschungsgemeinschaft.

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Correspondence to Albrecht Bindereif.

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Hui, J., Stangl, K., Lane, W. et al. HnRNP L stimulates splicing of the eNOS gene by binding to variable-length CA repeats. Nat Struct Mol Biol 10, 33–37 (2003). https://doi.org/10.1038/nsb875

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