p-120 Catenin is a Useful Diagnostic Biomarker for Distinguishing Plasmacytoid and Sarcomatoid Variants From Conventional Urothelial Carcinoma

Context.— Plasmacytoid urothelial carcinoma (PC-UC) is an aggressive variant of urothelial carcinoma (UC), characterized by loss of E-cadherin (E-Cad)–mediated intercellular adhesion. Loss of E-Cad by immunohistochemistry can help diagnosis PC-UC; however, sensitivity is limited. Expression of other cadherin-catenin adhesion complex members, that is, p-120 catenin (p-120) and β-catenin (B-Cat), which are diagnostically useful for lobular breast carcinoma, remains unknown in UC. Objective.— To determine the utility of p-120 and B-Cat in conventional and variant UC. Design.— E-cadherin, B-Cat, and p-120 immunohistochemistry was performed in 25 conventional UCs and 33 variant UCs, including 22 PC-UCs, 6 sarcomatoid UCs (SUCs), and 5 micropapillary UCs. Membranous staining for all biomarkers was considered normal; however, any cytoplasmic staining or an absence of staining was considered diagnostically abnormal. Next-generation sequencing was performed on 8 PC-UC cases. Results.— E-cadherin, B-Cat, and p-120 showed membranous staining in all conventional and micropapillary UCs. In contrast, most PC-UCs were negative for E-Cad (17 of 22; 77%) with an additional 2 of 22 cases (9%) showing cytoplasmic with partial membranous staining. p-120 catenin demonstrated cytoplasmic or negative staining in 21 of 22 cases (95%). Most SUCs showed an absence of E-Cad (5 of 6; 83%) and cytoplasmic or negative p-120 in 5 of 6 cases (83%). Staining for B-Cat was also abnormal in a subset of PC-UCs and SUCs. Five PC-UC cases that harbored CDH1 gene variants were p-120 cytoplasmic positive. Conclusions.— p-120 catenin is a useful adjunct biomarker to E-Cad in the clinically important distinction of PC-UC and SUC from conventional UC. In particular, the combination of cytoplasmic p-120 and loss of E-Cad is strongly supportive of PC-UC and SUC.

USP9X-mediated deubiquitination of B-cell CLL/lymphoma 9 potentiates Wnt signaling and promotes breast carcinogenesis

Hyperactivation of the canonical Wnt-signaling pathway is a prominent feature of a number of human malignancies. Transcriptional activation of this signaling cascade depends on the formation of the β-catenin–B-cell CLL/lymphoma 9 (BCL9)–pygopus (PYGO) family plant homeodomain finger 1 complex, yet how the assembly of this complex is regulated remains to be investigated. Here, using MCF-7, HeLa, HEK293T, MDA–MB-231, and Sf9 cells, along with immunoblotting and immunofluorescence, nano-HPLC–MS/MS, deubiquitination, immunoprecipitation, and chromatin immunoprecipitation (ChIP) assays, we report that BCL9 physically associates with a protein deubiquitinase, ubiquitin-specific peptidase 9, X-linked (USP9X), and that USP9X removes Lys-63–linked polyubiquitin on Lys-212 of BCL9. Importantly, the USP9X-mediated BCL9 deubiquitination facilitated the formation of the β-catenin–BCL9–PYGO complex, thereby potentiating the transcriptional activation of Wnt/β-catenin target genes. We also show that USP9X-mediated BCL9 deubiquitination promotes the proliferation and invasion of breast cancer cells. Together, these results uncover USP9X as a deubiquitinase of BCL9, implicating USP9X in Wnt/β-catenin signaling and breast carcinogenesis.

Increased rounds of gonadotropin stimulation have side effects on mouse fallopian tubes and oocytes

In this study, it was evaluated if increased rounds of gonadotropin stimulation could affect in mice: (i) expression levels of proteins regulating cell cycle and DNA repair in fallopian tubes and (ii) meiotic spindle morphology of ovulated oocytes. To this end, adult female mice were subjected or not (Control) to 6 or 8 rounds of gonadotropin stimulation. Ovulated oocytes were incubated with anti A/B tubulin to evaluate spindle morphology. Fallopian tubes were analyzed to detect Cyclin D1, phospho-p53/p53, phospho-AKT/AKT, phospho-GSK3B/GSK3B, SOX2, OCT3/4, phospho-B-catenin/B-catenin, phospho-CHK1 and phospho-H2A.X protein levels. After 6 rounds, Cyclin D1, p53 and phospho-p53 contents were higher than Control. After 8 rounds, the contents of phosphorylated AKT, GSK3B and p53 as well as of total p53, Cyclin D1 and OCT3/4 significantly increased in comparison with Control. Conversely, SOX2 and B-catenin were similarly expressed among all experimental groups. The finding that phospho-CHK1 and phospho-H2A.X protein levels were undetectable supported the absence of extensive DNA damage. Oocytes number and percentage of normal meiotic spindles drastically decreased from 6 rounds onward. Altogether, our results demonstrated that 6 and 8 cycles of gonadotropin stimulation reduce mouse reproductive performances by inducing over-expression and over-activation of proteins controlling cell cycle progression in fallopian tubes and by impairing oocyte spindle.