Alternative splicing and nonsense-mediated decay regulate telomerase reverse transcriptase (TERT) expression during virus-induced lymphomagenesis in vivo.
BACKGROUND
Telomerase activation, a critical step in cell immortalization and oncogenesis, is partly regulated by alternative splicing. In this study, we aimed to use the Marek's disease virus (MDV) T-cell lymphoma model to evaluate TERT regulation by splicing during lymphomagenesis in vivo, from the start point to tumor establishment.


RESULTS
We first screened cDNA libraries from the chicken MDV lymphoma-derived MSB-1 T- cell line, which we compared with B (DT40) and hepatocyte (LMH) cell lines. The chTERT splicing pattern was cell line-specific, despite similar high levels of telomerase activity. We identified 27 alternative transcripts of chicken TERT (chTERT). Five were in-frame alternative transcripts without in vitro telomerase activity in the presence of viral or chicken telomerase RNA (vTR or chTR), unlike the full-length transcript. Nineteen of the 22 transcripts with a premature termination codon (PTC) harbored a PTC more than 50 nucleotides upstream from the 3' splice junction, and were therefore predicted targets for nonsense-mediated decay (NMD). The major PTC-containing alternatively spliced form identified in MSB1 (ie10) was targeted to the NMD pathway, as demonstrated by UPF1 silencing. We then studied three splicing events separately, and the balance between in-frame alternative splice variants (d5f and d10f) plus the NMD target i10ec and constitutively spliced chTERT transcripts during lymphomagenesis induced by MDV indicated that basal telomerase activity in normal T cells was associated with a high proportion of in-frame non functional isoforms and a low proportion of constitutively spliced chTERT. Telomerase upregulation depended on an increase in active constitutively spliced chTERT levels and coincided with a switch in alternative splicing from an in-frame variant to NMD-targeted variants.


CONCLUSIONS
TERT regulation by splicing plays a key role in telomerase upregulation during lymphomagenesis, through the sophisticated control of constitutive and alternative splicing. Using the MDV T-cell lymphoma model, we identified a chTERT splice variant as a new NMD target.
Equipe TLVI, Université François Rabelais de Tours, UFR Sciences et Techniques, Parc de Grandmont 37200 Tours, France.
Souheila
S
Amor
Sylvie
S
Remy
Ginette
G
Dambrine
Yves
Y
Le Vern
Denis
D
Rasschaert
Sylvie
S
Laurent
D017398:Alternative Splicing; D000818:Animals; D002460:Cell Line; D002645:Chickens; D018076:DNA, Complementary / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D015723:Gene Library; D014644:Genetic Variation; D006801:Humans; D008223:Lymphoma / Q000201:enzymology* / Q000235:genetics* / Q000821:virology*; D019098:Telomerase / Q000096:biosynthesis* / Q000235:genetics*; D014162:Transfection; D015854:Up-Regulation
D018076:DNA, Complementary; D019098:Telomerase