Characterization of the role of Thoc1 in mouse development and tumorigenesis
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Regulation of transcription is integrated into the development and tumorigenesis of mammals. Transcription can be divided into several steps: promoter assembly, clearance and escape, elongation and termination. Recently Darzacq et al reported that polymerases enter a paused state for unexpectedly long times (Darzacq, Shav-Tal et al. 2007). Muse et al identified hundreds of genes with stalled RNA polymerases within the promoter-proximal region. Nearly one third of those genes with stalled RNA polymerases are involved in development. Genes involved in cell differentiation and cell communication are also enriched (Muse, Gilchrist et al. 2007). These findings indicate that regulation of transcriptional elongation plays an important role in development and homeostasis. Thoc1 functions in transcription elongation by participating in the TREX complex. TREX binds RNA polymerase II and couples transcriptional elongation to factors important for splicing and nuclear export. Recently we and others demonstrated that Thoc1 plays a unique role in tumor growth. Tumor cells express higher levels of Thoc1 and are more sensitive to loss of Thoc1 compared to normal cells (Li, Lin et al. 2007). However, the physiological role of Thoc1 in mammals is still unknown. Deletion of Hpr1p, the counterpart of Thoc1 in yeast, leads to temperature sensitive growth, hyperrecombination, increased sensitivity to genotoxic stress, xv defects in transcription, and nuclear export defects. Despite these defects, Hpr1p is not essential for viability in yeast. To address the physiological function of Thoc1, we constructed a Thoc1 knockout mouse. While heterozygous null Thoc1 mice are born at the expected Mendelian frequency, homozygous null mice are not recovered, indicating that Thoc1 is required for embryonic development. Embryonic development is arrested around the time of implantation. Cells of the inner cell mass, which is comprised of the pluripotent embryonic stem cells, are particularly dependent on Thoc1. Since early embryonic lethality precludes analysis of requirements for Thoc1 later in mouse development and in tumorigenesis, we have constructed a hypomorphic allele (H allele) and a conditional null allele (F allele). The hypomorphic allele expresses decreased levels of Thoc1 in all tissues examined. Mice homozygous for the hypomorphic allele are viable. However, they exhibit a runted phenotype. The relatively small size of Thoc1 H/H mice was detectable as early as embryonic day 12.5 (E12.5), suggesting the runted phenotype was not solely due to changes in the activity of growth factors and hormones that function during postnatal development. We also find that Thoc1 hypomorphic mice display dramatic defects in the development and function of the reproductive system. Male Thoc1 H/H mice are infertile. The testes are small with a severe deficiency in the number of mature sperm. Histology data demonstrates apoptosis in spermatocytes and differentiation defects in Sertoli cells. To further explore the gene expression patterns associated with the male infertility phenotype, mouse gene expression profiling has been conducted with 8 day old testes of Thoc1 H/H and wild-type littermate control mice. Consistent with the histology data, several Spermatogonia and Sertoli cell markers were down regulated, including c-kit, c-kit ligand, Ppt1, Rhox5, Gata1, AMHRII, Dhh, Sox9, Inhibitin-α and Nr5a1. This suggests a developmental defect in Spermatogonia and Sertoli cells of Thoc1 H/H mice. Thoc1 H/H female mice are subfertile, in part because of reduced numbers of follicles. These in vivo findings suggest a critical role for Thoc1 in growth and development of the reproductive system. Guo et al showed Thoc1 expression correlates with breast tumor size and metastatic potential (Guo, Hakimi et al. 2005). To further explore the role of Thoc1 in mammary gland tumorigenesis, we conditionally deleted Thoc1 in the mammary glands of the polyoma virus middle T transgenic mouse (MMTV-PyMT) model of breast cancer. Although we did not detect an obvious difference in tumor initiation and progression, we found that all resulting tumors retained expression of Thoc1, despite efficient Cre-mediated recombination in the mammary gland. Thus some MMTV-PyMT expressing mammary epithelial cells escape Cre-mediated Thoc1 excision, and such cells preferentially give rise to subsequent mammary tumors. This data is consistent with our recent unpublished data that conditional loss of Thoc1 in the prostate inhibits prostate tumor progression induced by p53 and Rb deficiency. Thoc1 expression may also be required for breast tumorigenesis in the mouse.