To evaluate the role of oncogenic RAS mutations in pancreatic tumorigenesis, we directed endogenous expression of KRAS(G12D) to progenitor cells of the mouse pancreas. We find that physiological levels of Kras(G12D) induce ductal lesions that recapitulate the full spectrum of human pancreatic intraepithelial neoplasias (PanINs), putative precursors to invasive pancreatic cancer. The PanINs are highly proliferative, show evidence of histological progression, and activate signaling pathways normally quiescent in ductal epithelium, suggesting potential therapeutic and chemopreventive targets for the cognate human condition. At low frequency, these lesions also progress spontaneously to invasive and metastatic adenocarcinomas, establishing PanINs as definitive precursors to the invasive disease. Finally, mice with PanINs have an identifiable serum proteomic signature, suggesting a means of detecting the preinvasive state in patients.

Erratum Preinvasive and invasive ductal pancreatic cancer and its early detection in the mouseIn the article by Hingorani et al. (Cancer Cell 4,, there are several typographical errors in the text regarding the citation of the figures. On page 441, the sentence "In many of the older mice, the pancreata contained extensive ductal lesions, and the acinar parenchyma was largely replaced by an intense stromal, or desmoplastic, reaction comprised of inflammatory cells, fibroblasts, and collagen deposition (Figures 2I-2K)" should instead refer to Figures 2I-2L. Also on page 441, the sentence "Finally, we note that PanINs expressed only low levels of PDX-1, which can nevertheless be discerned when compared to the lack of expression in surrounding acini (Figures 2G and 2H) and to normal ducts in control animals (Figure 2I)" should refer instead to Figure 3, and thus should read "Finally, we note that PanINs expressed only low levels of PDX-1, which can nevertheless be discerned when compared to the lack of expression in surrounding acini (Figures 3G and 3H) and to normal ducts in control animals (Figure 3I)."

Serum potentially carries an archive of important histological information whose determination could serve to improve early disease detection. The analysis of serum, however, is analytically challenging due to the high dynamic concentration range of constituent protein/peptide species, necessitating extensive fractionation prior to mass spectrometric analyses. The low molecular weight (LMW) serum proteome is that protein/peptide fraction from which high molecular weight proteins, such as albumin, immunoglobulins, transferrin, and lipoproteins, have been removed. This LMW fraction is made up of several classes of physiologically important proteins such as cytokines, chemokines, peptide hormones, as well as proteolytic fragments of larger proteins. Centrifugal ultrafiltration of serum was used to remove the large constituent proteins resulting in the enrichment of the LMW proteins/ peptides. Because albumin is known to bind and transport small molecules and peptides within the circulatory system, the centrifugal ultrafiltration was conducted under solvent conditions effecting the disruption of protein-protein interactions. The LMW serum proteome sample was digested with trypsin, fractionated by strong cation exchange chromatography, and analyzed by microcapillary reversed-phase liquid chromatography coupled on-line with electrospray ionization tandem mass spectrometry. Analysis of the tandem mass spectra resulted in the identification of over 340 human serum proteins; however, not a single peptide from serum albumin was observed. The large number of proteins identified demonstrates the efficacy of this method for the removal of large abundant proteins and the enrichment of the LMW serum proteome.

Stem cells are controlled, in part, by genetic pathways frequently dysregulated during human tumorigenesis. Either stimulation of Wnt/β-catenin signalling or overexpression of telomerase is sufficient to activate quiescent epidermal stem cells in vivo, although the mechanisms by which telomerase exerts these effects are not understood. Here we show that telomerase directly modulates Wnt/β-catenin signalling by serving as a cofactor in a β-catenin transcriptional complex. The telomerase protein component TERT (telomerase reverse transcriptase) interacts with BRG1 (also called SMARCA4), a SWI/SNF-related chromatin remodelling protein, and activates Wnt-dependent reporters in cultured cells and in vivo. TERT serves an essential role in formation of the anterior–posterior axis in Xenopus laevis embryos, and this defect in Wnt signalling manifests as homeotic transformations in the vertebrae of Tert−/− mice. Chromatin immunoprecipitation of the endogenous TERT protein from mouse gastrointestinal tract shows that TERT physically occupies gene promoters of Wnt-dependent genes. These data reveal an unanticipated role for telomerase as a transcriptional modulator of the Wnt/ β-catenin signalling pathway.

Infection with human immunodeficiency virus 1 (HIV-1) results in the dissemination of virus to gut-associated lymphoid tissue. Subsequently, HIV-1 mediates massive depletion of gut CD4+ T cells, which contributes to HIV-1-induced immune dysfunction. The migration of lymphocytes to gut-associated lymphoid tissue is mediated by integrin alpha4beta7. We demonstrate here that the HIV-1 envelope protein gp120 bound to an activated form of alpha4beta7. This interaction was mediated by a tripeptide in the V2 loop of gp120, a peptide motif that mimics structures presented by the natural ligands of alpha4beta7. On CD4+ T cells, engagement of alpha4beta7 by gp120 resulted in rapid activation of LFA-1, the central integrin involved in the establishment of virological synapses, which facilitate efficient cell-to-cell spreading of HIV-1.

Covalent modifications of histones, such as acetylation and methylation, play important roles in the regulation of gene expression. Histone lysine methylation has been implicated in both gene activation and repression, depending on the specific lysine (K) residue that becomes methylated and the state of methylation (mono-, di-, or trimethylation). Methylation on K4, K9, and K36 of histone H3 has been shown to be reversible and can be removed by site-specific demethylases. However, the enzymes that antagonize methylation on K27 of histone H3 (H3K27), an epigenetic mark important for embryonic stem cell maintenance, Polycombmediated gene silencing, and X chromosome inactivation have been elusive. Here we show the JmjC domain-containing protein UTX (ubiquitously transcribed tetratricopeptide repeat, X chromosome), as well as the related JMJD3 (jumonji domain containing 3), specifically removes methyl marks on H3K27 in vitro. Further, the demethylase activity of UTX requires a catalytically active JmjC domain. Finally, overexpression of UTX and JMJD3 leads to reduced di-and trimethylation on H3K27 in cells, suggesting that UTX and JMJD3 may function as H3K27 demethylases in vivo. The identification of UTX and JMJD3 as H3K27-specific demethylases provides direct evidence to indicate that similar to methylation on K4, K9, and K36 of histone H3, methylation on H3K27 is also reversible and can be dynamically regulated by site-specific histone methyltransferases and demethylases.histone methylation ͉ transcriptional regulation

Telomerase is a ribonucleoprotein (RNP) complex that synthesizes telomere repeats in tissue progenitor cells and cancer cells. Active human telomerase consists of at least three principal subunits, including the telomerase reverse transcriptase (TERT), the telomerase RNA (TERC), and dyskerin. Here, we identify a holoenzyme subunit, TCAB1 (telomerase Cajal body protein1), uniquely enriched in Cajal bodies, nuclear sites of RNP processing important for telomerase function. TCAB1 associates with active telomerase enzyme, with established telomerase components, and with small Cajal body RNAs involved in modifying splicing RNAs. Depletion of TCAB1 using RNA interference prevents TERC from associating with Cajal bodies, disrupts telomerase-telomere association and abrogates telomere synthesis by telomerase. Thus, TCAB1 controls telomerase trafficking and is required for telomere synthesis in human cancer cells.TERT and TERC comprise the minimal catalytic core of the telomerase enzyme (1), whereas dyskerin is an RNA binding protein that recognizes the H/ACA sequence motif shared by TERC and two groups of non-coding RNAs involved in RNA modification -small Cajal body (sca) RNAs and small nucleolar (sno) RNAs (2,3). Dyskerin functions in part to support telomerase RNP biogenesis and TERC stability (4,5). TERT, TERC and dyskerin are all components of active telomerase (6), and mutations in any of these genes can cause the human stem cell disorder dyskeratosis congenita (7). Other potential components of active telomerase include three evolutionarily conserved dyskerin-associated proteins, NOP10, NHP2 and GAR1 (8-10), and EST1A, a homologue of the yeast telomerase protein Est1p (11,12). However, the size of active human telomerase, estimated in the 0.65 to 2 MDa range (6,13,14), suggests the existence of additional components. We reasoned that other dyskerin-associated proteins may be telomerase components, and we therefore sought to purify dyskerin complexes.To study dyskerin, we expressed tagged dyskerin protein at endogenous levels in the absence of competing endogenous protein (Fig. S1) and isolated dyskerin complexes using a dual affinity chromatography strategy. Purified dyskerin complexes were analyzed by SDS-PAGE and nanoLC-MS/MS for identification of co-purifying proteins (Fig. 1A,B). Dense peptide coverage was obtained for dyskerin and for the dyskerin-associated ATPases pontin and reptin (14). Each of the evolutionarily conserved dyskerin-binding proteins NHP2, NOP10 and GAR1 was detected, as were the dyskerin-associated proteins Nopp140 and NAF1, a nucleoplasmic factor required for assembly of H/ACA RNPs including telomerase (15) (Fig. 1B). In addition, this approach identified the WD40 repeat protein WDR79, a protein that had not been previously implicated in dyskerin or telomerase function.We further characterized WDR79, hereafter referred to as TCAB1 (Fig. 1B, S2). Endogenous TCAB1 was specifically bound to Flag-dyskerin immunoprecipitated from Flagdyskerin +shRNA HeLa cells, as were endogenous ponti...

Summary Members of sirtuin family regulate multiple critical biological processes, yet their role in carcinogenesis remains controversial. To investigate the physiological functions of SIRT2 in development and tumorigenesis, we disrupted Sirt2 in mice. We demonstrated that SIRT2 regulates the anaphase-promoting complex/cyclosome activity through deacetylation of its co-activators, APCCDH1 and CDC20. SIRT2 deficiency caused increased levels of mitotic regulators, including Aurora-A and -B that direct centrosome amplification, aneuploidy, and mitotic cell death. Sirt2-deficient mice develop gender-specific tumorigenesis, with females primarily developing mammary tumors, and males developing more hepatocellular carcinoma (HCC). Human breast cancers and HCC samples exhibited reduced SIRT2 levels compared with normal tissues. These data demonstrate that SIRT2 is a tumor suppressor through its role in regulating mitosis and genome integrity.