Genomic imprinting at the WT1 gene involves a novel coding transcript (AWT1) that shows deregulation in Wilms' tumours

Abstract

The Wilms' tumour suppressor gene, WT1, is mutated in 10-15% of Wilms' tumours and encodes zinc-finger proteins with diverse cellular functions critical for nephrogenesis, genitourinary development, haematopoiesis and sex determination. Here we report that a novel alternative WT1 transcript, AWT1, is co-expressed with WT1 in renal and haematopoietic cells. AWT1 maintains WT1 exonic structure between exons 2 and 10, but deploys a new 5'-exon located in intron 1 of WT1. The AWT1 gene predicts proteins of approximately 33 kDa, comprising all exon 5 and exon 9 splicing variants previously characterized for WT1. Although WT1 is not genomically imprinted in kidney, we have previously shown monoallelic expression of a WT1 antisense transcript (WT1-AS) that is consistent with genomic imprinting. Here we demonstrate that both WT1-AS and the novel AWT1 transcript are imprinted in normal kidney with expression confined to the paternal allele. Wilms' tumours display biallelic AWT1 expression, indicating relaxation of imprinting of AWT1 in a subset of WTs. Our findings define human chromosome 11p13 as a new imprinted locus, and also suggest a possible molecular basis for the strong bias of paternal allele mutations and variable penetrance observed in syndromes with inherited WT1 mutations.

Frequency and Timing of Loss of Imprinting at 11p13 and 11p15 in Wilms' Tumor Development

“…We have previously shown that the WT1 locus at 11p13 expresses two imprinted transcripts: the noncoding antisense RNA WT1-AS (21) and the alternate coding transcript AWT1 (22). Hypomethylation of the DMR in the WT1 ARR is associated with LOI of the WT1-AS and AWT1 transcripts (21,22,25).…”

“…These imprinted transcripts are the noncoding antisense RNA WT1-AS (21) and the alternate coding transcript AWT1, which gives rise to NH 2 -terminally truncated WT1 proteins (22). We have shown that in a subset of WTs, LOI of these imprinted WT1 transcripts occurs, leading to their biallelic expression (21,22).…”

Real-time PCR quantification of major Wilms’ tumor gene 1 (WT1) isoforms in acute myeloid leukemia, their characteristic expression patterns and possible functional consequences

“…3,[10][11][12] At least 36 isoforms of WT1 protein are produced from the same DNA template as a result of alternative transcription initiation, alternative pre-mRNA splicing, mRNA editing and alternative translation initiation. 1,3,5,10,[13][14][15][16][17][18] The four major WT1 isoforms are generated by an alternative splicing and vary in the presence or absence of so called 17AA insert (17 amino acids encoded by the whole exon 5) and KTS insert (according to the three amino acids leucinethreonine-serine encoded by the terminal sequence of exon 9)- [19][20][21][22][23] An alternative WT1 transcript, AWT1 (also short transcript, sWT1), maintains WT1 exonic structure between exons 2 and 10 but uses another first exon located in intron 1 of WT1 (exon 1a). It may otherwise exist in all splicing variants characteristic of WT1.…”

“…It may otherwise exist in all splicing variants characteristic of WT1. 14,15 Another N-terminally truncated transcript, lacking exons 1-5, has been previously described and the presence of a cryptic promoter within WT1 intron 5 hypothesized. 24,25 All of these modifications markedly influence the activity of WT1 protein.…”

“…The AWT1 transcript does not contain the repression domain ( Figure 1). 1,3,14,15,26,27 Isoforms containing both splice inserts are the prevalent variants, the least common transcripts are those missing both inserts, and the physiological ratio of A[…”