Tissue-specific alternative splicing in the human INK4a/ARF cell cycle regulatory locus

The INK4a/ARF locus on human chromosome 9p resides at the nexus of two critical cell cycle regulatory pathways, the p53 pathway and the retinoblastoma (pRb) gene pathway. Through the use of shared coding regions and alternative reading frames two distinct proteins are produced: INK4a is a cyclin-dependent kinase inhibitor whereas ARF binds the MDM2 protooncogene and stabilizes p53. We have examined the expression patterns of the INK4a/ARF locus at the RNA level in normal human and murine tissues to determine if these genes are coordinately regulated. We found that both INK4a and ARF were expressed in most tissues at low levels detectable only by RT-PCR. The pancreas was an exception in that it expressed no detectable ARF mRNA but expressed high levels of INK4a mRNA. Furthermore, human pancreas expressed an additional previously unrecognized splice variant of INK4a, termed p12, through the use of an alternative splice donor site within intron 1. The p12 transcript produced a 12 kD protein composed of INK4a exon 1x and a novel intron-derived C-terminus. This novel protein did not interact with cdk4 but was capable of suppressing growth in a pRb-dependent manner. The implications of the capacity of the INK4a/ARF locus to encode a third transcript, and for pancreatic cancer, in which the INK4a/ARF locus is nearly always altered, are considered.