The retinoids are reported to reduce incidence of second primary aerodigestive cancers. Mechanisms for this chemoprevention are previously linked to all-trans retinoic acid (RA) signaling growth inhibition at G 1 in carcinogen-exposed immortalized human bronchial epithelial cells. This study investigated how RA suppresses human bronchial epithelial cell growth at the G 1 -S cell cycle transition. RA signaled growth suppression of human bronchial epithelial cells and a decline in cyclin D1 protein but not mRNA expression. Exogenous cyclin D1 protein also declined after RA treatment of transfected, immortalized human bronchial epithelial cells, suggesting that posttranslational mechanisms were active in this regulation of cyclin D1 expression. Findings were extended by showing treatment with ubiquitindependent proteasome inhibitors: calpain inhibitor I and lactacystin each prevented this decreased cyclin D1 protein expression, despite RA treatment. Treatment with the cysteine proteinase inhibitor, E-64, did not prevent this cyclin D1 decline. High molecular weight cyclin D1 protein species appeared after proteasome inhibitor treatments, suggesting that ubiquitinated species were present. To learn whether RA directly promoted degradation of cyclin D1 protein, studies using human bronchial epithelial cell protein extracts and in vitro-translated cyclin D1 were performed. In vitro-translated cyclin D1 degraded more rapidly when incubated with extracts from RA treated vs. untreated cells. Notably, this RA-signaled cyclin D1 proteolysis depended on the C-terminal PEST sequence, a region rich in proline (P), glutamate (E), serine (S), and threonine (T). Taken together, these data highlight RA-induced cyclin D1 proteolysis as a mechanism signaling growth inhibition at G 1 active in the prevention of human bronchial epithelial cell transformation.

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All-trans-retinoic acid (RA) treatment induces remissions in acute promyelocytic leukemia (APL) cases expressing the t(15;17) product, promyelocytic leukemia (PML)͞RA receptor ␣ (RAR␣). Microarray analyses previously revealed induction of UBE1L (ubiquitin-activating enzyme E1-like) after RA treatment of NB4 APL cells. We report here that this occurs within 3 h in RA-sensitive but not RA-resistant APL cells, implicating UBE1L as a direct retinoid target. A 1.3-kb fragment of the UBE1L promoter was capable of mediating transcriptional response to RA in a retinoid receptor-selective manner. PML͞RAR␣, a repressor of RA target genes, abolished this UBE1L promoter activity. A hallmark of retinoid response in APL is the proteasome-dependent PML͞RAR␣ degradation. UBE1L transfection triggered PML͞RAR␣ degradation, but transfection of a truncated UBE1L or E1 did not cause this degradation. A tight link was shown between UBE1L induction and PML͞RAR␣ degradation. Notably, retroviral expression of UBE1L rapidly induced apoptosis in NB4 APL cells, but not in cells lacking PML͞RAR␣ expression. UBE1L has been implicated directly in retinoid effects in APL and may be targeted for repression by PML͞ RAR␣. UBE1L is proposed as a direct pharmacological target that overcomes oncogenic effects of PML͞RAR␣ by triggering its degradation and signaling apoptosis in APL cells.