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Volume 272, Number 52, Issue of December 26, 1997 pp. 33394-33401

Novel Exonic Elements That Modulate Splicing of the Human Fibronectin EDA Exon

(Received for publication, August 20, 1997, and in revised form, October 21, 1997)

Alfredo Staffa , Nicholas H. Acheson and Alan Cochrane ^¶

From the  Department of Microbiology and Immunology, McGill University, Montreal, Quebec H3A 2B4, and the ^¶ Department of Medical Genetics and Microbiology, University of Toronto, Toronto, Ontario M5S 1A8, Canada

Three exons in the fibronectin primary transcript are alternatively spliced in a tissue- and developmental stage-specific manner. One of these exons, EDA, has been shown previously by others to contain two splicing regulatory elements between 155 and 180 nucleotides downstream of the 3-splice site: an exon splicing enhancer and a negative element. By transient expression of a chimeric -globin/fibronectin EDA intron in COS-7 cells, we have identified two additional exonic splicing regulatory elements. RNA generated by a construct containing the first 120 nucleotides of the fibronectin EDA exon was spliced with an efficiency of approximately 50%. Deletion of most of the fibronectin EDA exon sequences resulted in a 20-fold increase in the amount of spliced RNA, indicative of an exon splicing silencer. Deletion and mutagenesis studies suggest that the fibronectin exon splicing silencer is associated with a conserved RNA secondary structure. In addition, sequences between nucleotides 93 and 118 of the EDA exon contain a non-purine-rich splicing enhancer as demonstrated by its ability to function in a heterologous context.

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