Loss of CDKN2A expression is a frequent event in primary invasive melanoma and correlates with sensitivity to the CDK4/6 inhibitor PD0332991 in melanoma cell lines

We have investigated the potential for the p16‐cyclin D‐CDK4/6‐retinoblastoma protein pathway to be exploited as a therapeutic target in melanoma. In a cohort of 143 patients with primary invasive melanoma, we used fluorescence in situ hybridization to detect gene copy number variations (CNVs) in CDK4, CCND1, and CDKN2A and immunohistochemistry to determine protein expression. CNVs were common in melanoma, with gain of CDK4 or CCND1 in 37 and 18% of cases, respectively, and hemizygous or homozygous loss of CDKN2A in 56%. Three‐quarters of all patients demonstrated a CNV in at least one of the three genes. The combination of CCND1 gain with either a gain of CDK4 and/or loss of CDKN2A was associated with poorer melanoma‐specific survival. In 47 melanoma cell lines homozygous loss, methylation or mutation of CDKN2A gene or loss of protein (p16^INK4A) predicted sensitivity to the CDK4/6 inhibitor PD0332991, while RB1 loss predicted resistance.     

Cooperation between the chloroplast psbA 5′‐untranslated region and coding region is important for translational initiation: the chloroplast translation machinery cannot read a human viral gene coding region

Chloroplast mRNA translation is regulated by the 5′‐untranslated region (5′‐UTR). Chloroplast 5′‐UTRs also support translation of the coding regions of heterologous genes. Using an in vitro translation system from tobacco chloroplasts, we detected no translation from a human immunodeficiency virus tat coding region fused directly to the tobacco chloroplast psbA 5′‐UTR. This lack of apparent translation could have been due to rapid degradation of mRNA templates or synthesized protein products. Replacing the psbA 5′‐UTR with the E. coli phage T7 gene 10 5′‐UTR, a highly active 5′‐UTR, and substituting synonymous codons led to some translation of the tat coding region. The Tat protein thus synthesized was stable during translation reactions. No significant degradation of the added tat mRNAs was observed after translation reactions. These results excluded the above two possibilities and confirmed that the tat coding region prevented its own translation. The tat coding region was then fused to the psbA 5′‐UTR with a cognate 5′‐coding segment. Significant translation was detected from the tat coding region when fused after 10 or more codons. That is, translation could be initiated from the tat coding region once translation had started, indicating that the tat coding region inhibits translational initiation but not elongation. Hence, cooperation/compatibility between the 5′‐UTR and its coding region is important for translational initiation.