SUMMARY Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and human pancreas tissues. Pancreatic organoids can be rapidly generated from resected tumors and biopsies, survive cryopreservation and exhibit ductal- and disease stage-specific characteristics. Orthotopically transplanted neoplastic organoids recapitulate the full spectrum of tumor development by forming early-grade neoplasms that progress to locally invasive and metastatic carcinomas. Due to their ability to be genetically manipulated, organoids are a platform to probe genetic cooperation. Comprehensive transcriptional and proteomic analyses of murine pancreatic organoids revealed genes and pathways altered during disease progression. The confirmation of many of these protein changes in human tissues demonstrates that organoids are a facile model system to discover characteristics of this deadly malignancy.

The heterogeneity and instability of human tumors hamper straightforward identification of cancer-causing mutations through genomic approaches alone. Herein we describe a mouse model of liver cancer initiated from progenitor cells harboring defined cancer-predisposing lesions. Genome-wide analyses of tumors in this mouse model and in human hepatocellular carcinomas revealed a recurrent amplification at mouse chromosome 9qA1, the syntenic region of human chromosome 11q22. Gene-expression analyses delineated cIAP1, a known inhibitor of apoptosis, and Yap, a transcription factor, as candidate oncogenes in the amplicon. In the genetic context of their amplification, both cIAP1 and Yap accelerated tumorigenesis and were required to sustain rapid growth of amplicon-containing tumors. Furthermore, cIAP1 and Yap cooperated to promote tumorigenesis. Our results establish a tractable model of liver cancer, identify two oncogenes that cooperate by virtue of their coamplification in the same genomic locus, and suggest an efficient strategy for the annotation of human cancer genes.

Programmed cell death (apoptosis) is an evolutionarily conserved process used by multicellular organisms to eliminate cells that are not needed or are potentially detrimental to the organism. Members of the Bcl-2 family of mammalian proteins are intimately involved in the regulation of apoptosis, but, their precise mechanism of action remains unresolved. In Caenorhabditis elegans, the Bcl-2 homologue CED-9 prevents cell death by antagonizing the death-promoting activities of CED-3, a member of the Caspase family of death proteases, and of CED-4, a protein with no known mammalian homologue. Here we show that CED-9 interacts physically with CED-4. Mutations that reduce or eliminate CED-9 activity also disrupt its ability to bind CED-4, suggesting that this interaction is important for CED-9 function. Thus, CED-9 might control C. elegans cell death by binding to and regulating CED-4 activity. We propose that mammalian Bcl-2 family members might control apoptosis in a similar way through interaction and regulation of CED-4 homologues or analogues.

The HIR/HPC (histone regulation/histone periodic control) negative regulators play important roles in the transcription of six of the eight core histone genes during the Saccharomyces cerevisiae cell cycle. The phenotypes of hir1 and hir2 mutants suggested that the wild-type HIR1 and HIR2 genes encode transcriptional repressors that function in the absence of direct DNA binding. When Hir1p and Hir2p were artificially tethered to yeast promoters, each protein repressed transcription, suggesting that they represent a new class of transcriptional corepressors. The two proteins might function as a complex in vivo: Hir2p required both Hir1p and another Hir protein, Hir3p, to repress transcription when it was tethered to an HTA1-lacZ reporter gene, and Hir1p and Hir2p could be coimmunoprecipitated from yeast cell extracts. Tethered Hir1p also directed the periodic transcription of the HTA1 gene and repressed HTA1 transcription in response to two cell cycle regulatory signals. Thus, it represents the first example of a transcriptional corepressor with a direct role in cell cycle-regulated transcription.Histone synthesis is tightly regulated in eukaryotes to ensure that the components for nucleosome assembly are present both at the correct time during the cell cycle and in the proper stoichiometric levels (39). Regulation at the level of transcription plays an important role in the synthesis of the core histones in Saccharomyces cerevisiae, and negative regulation has emerged as a significant mechanism in the transcription of six of the eight histone genes (HTA1-HTB1, HHT1-HHF1, and HHT2-HHF2) in this organism (39). A negative site has been identified in the promoters of each of the negatively regulated histone genes in close proximity to upstream activation sequence (UAS) elements (14, 41). The HTA1-HTB1 negative site has been shown to play a key role in cell cycle-regulated transcription: deletion of this site derepresses HTA1 and HTB1 transcription early in G 1 phase, late in S phase, and in G 2 /M phase, points in the cell cycle when the two genes are usually not expressed (41), and prevents the same genes from being repressed in S-phase cells when DNA replication has been interrupted (29). The same site is also required for a feedback repression pathway that is triggered by the overproduction of histones H2A and H2B (32). In its absence, HTA1 and HTB1 transcription is insensitive to the intracellular levels of these two histone proteins.Two related genetic screens have identified seven HIR and HPC (histone regulation and histone periodic control) genes whose products are postulated to encode repressors of HTA1-HTB1, HHT1-HHF1, and HHT2-HHF2 transcription (43, 64). Four of these genes (HIR1, HIR2, HIR3, and HPC2) have been isolated and extensively characterized (51,54,64). None of the genes is essential, and each appears to encode a novel polypeptide. Interestingly, the HIR1 and HIR2 genes have been evolutionarily conserved; the human homolog, HIRA, encodes a protein that encompasses structural features from both ye...