Yap/Taz are required for establishing the cerebellar radial glia scaffold and proper foliation.

Yap/Taz are well-established downstream effectors of the Hippo pathway, known to regulate organ size by directing proliferation and apoptosis. Although the functions of Yap/Taz have been extensively studied, little is known about their role in brain development. Here, through genetic ablation, we show that Yap/Taz are required for cerebellar morphogenesis. Yap/Taz deletion in neural progenitors causes defects in secondary fissure formation, leading to abnormal folia development. Although they seemed very likely to serve an important function in the development of cerebellar granule cell precursors, Yap/Taz are dispensable for their proliferation. Furthermore, Yap/Taz loss does not rescue the medulloblastoma phenotype caused by constitutively active Smoothened. Importantly, Yap/Taz are highly expressed in radial glia and play a crucial role in establishing the radial scaffold and cellular polarity of neural progenitors during embryogenesis. We found that Yap/Taz are necessary to establish and maintain junctional integrity of cerebellar neuroepithelium as prominent junction proteins are not maintained at the apical junction in the absence of Yap/Taz. Our study identifies a novel function of Yap/Taz in cerebellar foliation and finds that they are required to establish the radial glia scaffold and junctional stability.

Yap is required for ependymal integrity and is suppressed in LPA-induced hydrocephalus.

Timely generation and normal maturation of ependymal cells along the aqueduct are critical for preventing physical blockage between the third and fourth ventricles and the development of fetal non-communicating hydrocephalus. Our study identifies Yap, the downstream effector of the evolutionarily conserved Hippo pathway, as a central regulator for generating developmentally controlled ependymal cells along the ventricular lining of the aqueduct. Yap function is necessary for proper proliferation of progenitors and apical attachment of ependymal precursor cells. Importantly, an injury signal initiated by lysophosphatidic acid (LPA), an upstream regulator of Yap that can cause fetal haemorrhagic hydrocephalus, deregulates Yap in the developing aqueduct. LPA exposure leads to the loss of N-cadherin concentrations at the apical endfeet, which can be partially restored by forced Yap expression and more efficiently by phosphomimetic Yap. These results reveal a novel function of Yap in retaining tissue junctions during normal development and after fetal brain injury.

The apical complex protein Pals1 is required to maintain cerebellar progenitor cells in a proliferative state.

Through their biased localization and function within the cell, polarity complex proteins are necessary to establish the cellular asymmetry required for tissue organization. Well-characterized germinal zones, mitogenic signals and cell types make the cerebellum an excellent model for addressing the crucial function of polarity complex proteins in the generation and organization of neural tissues. Deletion of the apical polarity complex protein Pals1 in the developing cerebellum results in a remarkably undersized cerebellum with disrupted layers in poorly formed folia and strikingly reduced granule cell production. We demonstrate that Pals1 is not only essential for cerebellum organogenesis, but also for preventing premature differentiation and thus maintaining progenitor pools in cerebellar germinal zones, including cerebellar granule neuron precursors in the external granule layer. In the Pals1 mouse mutants, the expression of genes that regulate the cell cycle was diminished, correlating with the loss of the proliferating cell population of germinal zones. Furthermore, enhanced Shh signaling through activated Smo cannot overcome impaired cerebellar cell generation, arguing for an epistatic role of Pals1 in proliferation capacity. Our study identifies Pals1 as a novel intrinsic factor that regulates the generation of cerebellar cells and Pals1 deficiency as a potential inhibitor of overactive mitogenic signaling.

Impaired Reelin-Dab1 Signaling Contributes to Neuronal Migration Deficits of Tuberous Sclerosis Complex.

Tuberous sclerosis complex (TSC) is associated with neurodevelopmental abnormalities, including defects in neuronal migration. However, the alterations in cell signaling mechanisms critical for migration and final positioning of neurons in TSC remain unclear. Our detailed cellular analyses reveal that reduced Tsc2 in newborn neurons causes abnormalities in leading processes of migrating neurons, accompanied by significantly delayed migration. Importantly, we demonstrate that Reelin-Dab1 signaling is aberrantly regulated in TSC mouse models and in cortical tubers from TSC patients owing to enhanced expression of the E3 ubiquitin ligase Cul5, a known mediator of pDab1 ubiquitination. Likewise, mTORC1 activation by Rheb overexpression generates similar neuronal and Reelin-Dab1 signaling defects, and directly upregulates Cul5 expression. Inhibition of mTORC1 by rapamycin treatment or by reducing Cul5 largely restores normal leading processes and positioning of migrating neurons. Thus, disrupted Reelin-Dab1 signaling is critically involved in the neuronal migration defects of TSC.

Dual function of Yap in the regulation of lens progenitor cells and cellular polarity.

Hippo-Yap signaling has been implicated in organ size determination via its regulation of cell proliferation, growth and apoptosis (Pan, 2007). The vertebrate lens comprises only two major cell types, lens progenitors and differentiated fiber cells, thereby providing a relatively simple system for studying size-controlling mechanisms. In order to investigate the role of Hippo-Yap signaling in lens size regulation, we conditionally ablated Yap in the developing mouse lens. Lens progenitor-specific deletion of Yap led to near obliteration of the lens primarily due to hypocellularity in the lens epithelium (LE) and accompanying lens fiber (LF) defects. A significantly reduced LE progenitor pool resulted mainly from failed self-renewal and increased apoptosis. Additionally, Yap-deficient lens progenitor cells precociously exited the cell cycle and expressed the LF marker, ß-Crystallin. The mutant progenitor cells also exhibited multiple cellular and subcellular alterations including cell and nuclear shape change, organellar polarity disruption, and disorganized apical polarity complex and junction proteins such as Crumbs, Pals1, Par3 and ZO-1. Yap-deficient LF cells failed to anchor to the overlying LE layer, impairing their normal elongation and packaging. Furthermore, our localization study results suggest that, in the developing LE, Yap participates in the cell context-dependent transition from the proliferative to differentiation-competent state by integrating cell density information. Taken together, our results shed new light on Yap's indispensable and novel organizing role in mammalian organ size control by coordinating multiple events including cell proliferation, differentiation, and polarity.

Validation of association of genetic variants at 10q with prostate-specific antigen (PSA) levels in men at high risk for prostate cancer.

OBJECTIVE: To validate six previously identified markers among men at increased risk of prostate cancer (African-American men and those with a family history of prostate cancer) enrolled in the Prostate Cancer Risk Assessment Program (PRAP), a prostate cancer screening study. PATIENTS AND METHODS: Eligibility criteria for PRAP include age 35-69 years with a family history of prostate cancer, African-American ethnicity regardless of family history, and known BRCA gene mutations. The genome-wide association study markers assessed included rs2736098 (5p15.33), rs10993994 (10q11), rs10788160 (10q26), rs11067228 (12q24), rs4430796 (17q12) and rs17632542 (19q13.33). Genotyping methods included either the Taqman(®) single nucleotide polymorphism (SNP) genotyping assay (Applied Biosystems, Foster City, CA, USA) or pyrosequencing. Linear regression models were used to evaluate the association between individual markers and log-transformed baseline PSA levels, while adjusting for potential confounders. RESULTS: A total of 707 participants (37% Caucasian, 63% African-American) with clinical and genotype data were included in the analysis. Rs10788160 (10q26) was strongly associated with PSA levels among Caucasian participants in the high-risk group (P < 0.01), with a 33.2% increase in PSA level with each A-allele carried. Furthermore, rs10993994 (10q11) was found to be associated with PSA level (P = 0.03) in Caucasian men in the high-risk group, with a 15% increase in PSA level with each T-allele carried. A PSA adjustment model based on allele carrier status at rs10788160 and rs10993994 was proposed, specific to high-risk Caucasian men. CONCLUSIONS: Genetic variation at 10q may be particularly important in personalizing the interpretation of PSA level for Caucasian men in the high-risk group. Such information may have clinical relevance in shared decision-making and individualized prostate cancer screening strategies for Caucasian men in the high-risk group, although further study is warranted.

Whole exome sequence analysis of serous borderline tumors of the ovary.

OBJECTIVE: Serous borderline tumor (SBT) is a unique histopathologic entity of the ovary, believed to be intermediate between benign cystadenoma and invasive low-grade serous carcinoma. While somatic mutations in the KRAS or BRAF, and rarely ERBB2, genes have been well characterized in SBTs, other genetic alterations have not been described. Toward a more comprehensive understanding of the molecular genetic architecture of SBTs, we undertook whole exome sequencing of this tumor type. METHODS: Following pathologic review and laser capture microdissection to enrich for tumor cells, whole exomes were prepared from DNA of two independent SBTs and subjected to massively parallel DNA sequencing. RESULTS: Both tumors contained an activating mutation of the BRAF gene. A total of 15 additional somatic mutations were identified, nine in one tumor and six in the other. Eleven were missense mutations and four were nonsense or deletion mutations. Fourteen of the 16 genes found to be mutated in this study have been reported to be mutated in other cancers. Furthermore, 12 of these genes are mutated in ovarian cancers. The FBXW7 and KIAA1462 genes are noteworthy candidates for a pathogenic role in serous borderline tumorigenesis. CONCLUSIONS: These findings suggest that a very small number of somatic genetic mutations are characteristic of SBTs of the ovary, thus supporting their classification as a relatively genetically stable tumor type. The mutant genes described herein represent novel candidates for the pathogenesis of ovarian SBT.

Assessing the clinical role of genetic markers of early-onset prostate cancer among high-risk men enrolled in prostate cancer early detection.

BACKGROUND: Men with familial prostate cancer and African American men are at risk for developing prostate cancer at younger ages. Genetic markers predicting early-onset prostate cancer may provide clinically useful information to guide screening strategies for high-risk men. We evaluated clinical information from six polymorphisms associated with early-onset prostate cancer in a longitudinal cohort of high-risk men enrolled in prostate cancer early detection with significant African American participation. METHODS: Eligibility criteria include ages 35 to 69 with a family history of prostate cancer or African American race. Participants undergo screening and biopsy per study criteria. Six markers associated with early-onset prostate cancer [rs2171492 (7q32), rs6983561 (8q24), rs10993994 (10q11), rs4430796 (17q12), rs1799950 (17q21), and rs266849 (19q13)] were genotyped. Cox models were used to evaluate time to prostate cancer diagnosis and prostate-specific antigen (PSA) prediction for prostate cancer by genotype. Harrell's concordance index was used to evaluate predictive accuracy for prostate cancer by PSA and genetic markers. RESULTS: Four hundred and sixty participants with complete data and ≥ 1 follow-up visit were included. Fifty-six percent were African American. Among African American men, rs6983561 genotype was significantly associated with earlier time to prostate cancer diagnosis (P = 0.005) and influenced prediction for prostate cancer by the PSA (P < 0.001). When combined with PSA, rs6983561 improved predictive accuracy for prostate cancer compared with PSA alone among African American men (PSA = 0.57 vs. PSA + rs6983561 = 0.75, P = 0.03). CONCLUSIONS: Early-onset marker rs6983561 adds potentially useful clinical information for African American men undergoing prostate cancer risk assessment. Further study is warranted to validate these findings. IMPACT: Genetic markers of early-onset prostate cancer have potential to refine and personalize prostate cancer early detection for high-risk men.

Racial differences in prediction of time to prostate cancer diagnosis in a prospective screening cohort of high-risk men: effect of TMPRSS2 Met160Val.

INTRODUCTION: To evaluate the TMPRSS2-ERG gene polymorphism with respect to self-identified race or ethnicity (SIRE), time to prostate cancer (PCA) diagnosis, and screening parameters in the Prostate Cancer Risk Assessment Program, a prospective screening program for high-risk men. PATIENTS AND METHODS: A total of 631 men aged between 35 and 69 years were studied. 'High-risk' was defined as ≥ one first degree or two second degree relatives with PCA, any African American (AA) man regardless of familial PCA, and men with BRCA1/2 mutations. Men with elevated prostate-specific antigen (PSA) concentrations or other indications for PCA underwent biopsy. Men were followed from time of study entry to PCA diagnosis. Cox models were used to evaluate time to PCA diagnosis by genotype. RESULTS: Genotype distribution differed significantly by SIRE (CT/TT vs CC, P < 0.0001). Among 183 Caucasian men with at least one follow-up visit, PCA was more than doubled in men carrying CT/TT vs CC genotypes (hazard ratio = 2.55, 95% CI = 1.14-5.70) after controlling for age and PSA. No association was seen among AA men by TMPRSS2 genotype. CONCLUSIONS: The T-allele of the Met160Val variant in TMPRSS2, which has been associated with the TMPRSS2-ERG fusion, may be informative of time to PCA diagnosis for a subset of high-risk Caucasian men who are undergoing regular PCA screening. This variant, along with other genetic markers, warrant further study for personalizing PCA screening.