Probes for INS

Telomerase consists of at least two essential elements, an RNA component hTR or TERC that contains the template for telomere DNA addition, and a catalytic reverse transcriptase (TERT). While expression of TERT has been considered the key rate limiting component for telomerase activity, increasing evidence suggests an important role for the regulation of TERC in telomere maintenance and perhaps other functions in human cancer. By using three orthogonal methods including RNAseq, RT-qPCR, and an analytically validated chromogenic RNA in situ hybridization assay, we report consistent overexpression of TERC in prostate cancer. This overexpression occurs at the precursor stage (e.g. high grade prostatic intraepithelial neoplasia or PIN), and persists throughout all stages of disease progression. Levels of TERC correlate with levels of MYC (a known driver of prostate cancer) in clinical samples and we also show the following: forced reductions of MYC result in decreased TERC levels in 8 cancer cell lines (prostate, lung, breast, and colorectal); forced overexpression of MYC in PCa cell lines, and in the mouse prostate, results in increased TERC levels; human TERC promoter activity is decreased after MYC silencing; and MYC occupies the TERC locus as assessed by chromatin immunoprecipitation (ChIP). Finally, we show that knockdown of TERC by siRNA results in reduced proliferation of prostate cancer cell lines. These studies indicate that TERC is consistently overexpressed in all stages of prostatic adenocarcinoma, and its expression is regulated by MYC. These findings nominate TERC as a novel prostate cancer biomarker and therapeutic target.

Hepatocytes are replenished gradually during homeostasis and robustly after liver injury1, 2. In adults, new hepatocytes originate from the existing hepatocyte pool3-8, but the cellular source of renewing hepatocytes remains unclear. Telomerase is expressed in many stem cell populations, and mutations in telomerase pathway genes have been linked to liver diseases9-11. Here we identify a subset of hepatocytes that expresses high levels of telomerase and show that this hepatocyte subset repopulates the liver during homeostasis and injury. Using lineage tracing from the telomerase reverse transcriptase (Tert) locus in mice, we demonstrate that rare hepatocytes with high telomerase expression (TERTHigh hepatocytes) are distributed throughout the liver lobule. During homeostasis, these cells regenerate hepatocytes in all lobular zones, and both self-renew and differentiate to yield expanding hepatocyte clones that eventually dominate the liver. In response to injury, the repopulating activity of TERTHigh hepatocytes is accelerated and their progeny cross zonal boundaries. RNA sequencing shows that metabolic genes are downregulated in TERTHigh hepatocytes, indicating that metabolic activity and repopulating activity may be segregated within the hepatocyte lineage. Genetic ablation of TERTHigh hepatocytes combined with chemical injury causes a marked increase in stellate cell activation and fibrosis. These results provide support for a 'distributed model' of hepatocyte renewal in which a subset of hepatocytes dispersed throughout the lobule clonally expands to maintain liver mass.

Abstract

Ephrin type-A receptor 2 (EPHA2) and one of its ligands, ephrin-A5 (EFNA5), have been associated with loss of eye lens transparency, or cataract, - an important cause of visual impairment. Here we show that mice functionally lacking EPHA2 (Epha2-null), EFNA5(Efna5-null), or both receptor and ligand (Epha2/Efna5-null) consistently develop mostly transparent lenses with an internal refractive disturbance and a grossly disturbed cellular architecture. In situ hybridization localized Epha2 and Efna5 transcripts to lens epithelial cells and nascent fiber cells at the lens equator. In vivo labeling of Epha2-null lenses with a thymidine analog detected a significant decrease in lens epithelial cell proliferation within the germinative zone resulting in impaired early lens growth. Ex vivo imaging of Epha2-null, Efna5-null, and Epha2/Efna5-null lenses labelled in vivo with a membrane-targeted red fluorescent protein revealed misalignment of elongating fiber cells at the lens equator and loss of Y-suture pattern formation near the anterior and posterior poles of the lens. Immuno-fluorescent labeling of lens major intrinsic protein or aquaporin-0 (MIP/AQP0) showed that the precise, radial column patterning of hexagonal fiber cells throughout the cortex region was disrupted in Epha2-null, Efna5-null and Epha2/Efna5-null lenses. Collectively, these data suggest that Epha2 and Efna5 participate in the complex, global patterning of lens fiber cells that is necessary for maximal optical quality.