hnRNP L regulates the tumorigenic capacity of lung cancer xenografts in mice via caspase-9 pre-mRNA processing.

Caspase-9 is involved in the intrinsic apoptotic pathway and suggested to play a role as a tumor suppressor. Little is known about the mechanisms governing caspase-9 expression, but post-transcriptional pre-mRNA processing generates 2 splice variants from the caspase-9 gene, pro-apoptotic caspase-9a and anti-apoptotic caspase-9b. Here we demonstrate that the ratio of caspase-9 splice variants is dysregulated in non-small cell lung cancer (NSCLC) tumors. Mechanistic analysis revealed that an exonic splicing silencer (ESS) regulated caspase-9 pre-mRNA processing in NSCLC cells. Heterogeneous nuclear ribonucleoprotein L (hnRNP L) interacted with this ESS, and downregulation of hnRNP L expression induced an increase in the caspase-9a/9b ratio. Although expression of hnRNP L lowered the caspase-9a/9b ratio in NSCLC cells, expression of hnRNP L produced the opposite effect in non-transformed cells, suggesting a post-translational modification specific for NSCLC cells. Indeed, Ser52 was identified as a critical modification regulating the caspase-9a/9b ratio. Importantly, in a mouse xenograft model, downregulation of hnRNP L in NSCLC cells induced a complete loss of tumorigenic capacity that was due to the changes in caspase-9 pre-mRNA processing. This study therefore identifies a cancer-specific mechanism of hnRNP L phosphorylation and subsequent lowering of the caspase-9a/9b ratio, which is required for the tumorigenic capacity of NSCLC cells.

Growth factor-stimulated mitogen-activated kinase (MAPK) phosphorylation in the rat epididymis is limited by segmental boundaries.

Previous evidence has shown that sperm maturation is the result of successive events that influence sperm cells as they move through different microenvironments from the caput to the cauda epididymis. The physiological basis for the creation and maintenance of specific microenvironments along the epididymis are poorly understood. Anatomically, the epididymis consists of segments or lobules of epididymal tubule separated by connective tissue septa (CTS). The fact that CTS restrict the diffusion of tracer substances between segments and that certain gene expression patterns are segment-specific suggest that segments may represent functional epididymal units. In this report, we have further investigated epididymal segmentation by focusing on the ability of CTS to limit the effect of biologically relevant molecules, in particular epidermal growth factor (EGF), basic fibroblast growth factor (FGF2), and vascular endothelial growth factor A (VEGFA), in Segments 1 and 2 of the rat epididymis. We have demonstrated that these growth factors activate mitogen-activated kinase (MAPK) in both segments studied and that growth factors injected into the interstitial space of these segments in vivo exhibited a stimulatory effect only in the segment into which they were injected, i.e., MAPK activation was not observed in the adjacent segment. This restricting influence of CTS was abrogated by treatment with collagenase. In addition, we demonstrate the expression of selected forms of these growth factors and their receptors in Segments 1 and 2, and identify potential downstream targets. These results suggest that CTS regulate the trophic influences of growth factors and potentially other paracrine molecules, thus creating functionally separate units within the epididymis.