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The extent of somatic mutation and clonal selection in the human bladder remains unknown. We sequenced 2097 bladder microbiopsies from 20 individuals using targeted (n = 1914 microbiopsies), whole-exome (n = 655), and whole-genome (n = 88) sequencing. We found widespread positive selection in 17 genes. Chromatin remodeling genes were frequently mutated, whereas mutations were absent in several major bladder cancer genes. There was extensive interindividual variation in selection, with different driver genes dominating the clonal landscape across individuals. Mutational signatures were heterogeneous across clones and individuals, which suggests differential exposure to mutagens in the urine. Evidence of APOBEC mutagenesis was found in 22% of the microbiopsies. Sequencing multiple microbiopsies from five patients with bladder cancer enabled comparisons with cancer-free individuals and across histological features. This study reveals a rich landscape of mutational processes and selection in normal urothelium with large heterogeneity across clones and individuals.

Slu7 and Prp18 act in concert during the second step of yeast pre-mRNA splicing. Here we show that the 382-amino-acid Slu7 protein contains two functionally important domains: a zinc knuckle (122CRNCGEAGHKEKDC135) and a Prp18-interaction domain (215EIELMKLELY224). Alanine cluster mutations of 215EIE217 and 221LELY224 abrogated Slu7 binding to Prp18 in a two-hybrid assay and in vitro, and elicited temperature-sensitive growth phenotypes in vivo. Yet, the mutations had no impact on Slu7 function in pre-mRNA splicing in vitro. Single alanine mutations of zinc knuckle residues Cys122, His130, and Cys135 had no effect on cell growth, but caused Slu7 function during pre-mRNA splicing in vitro to become dependent on Prp18. Specifically, zinc knuckle mutants required Prp18 in order to bind to the spliceosome. Compound mutations in both Slu7 domains (e.g., C122A-EIE, H130A-EIE, and C135A-EIE) were lethal in vivo and abolished splicing in vitro, suggesting that the physical interaction between Slu7 and Prp18 is important for cooperation in splicing. Depletion/reconstitution studies coupled with immunoprecipitations suggest that second step factors are recruited to the spliceosome in the following order: Slu7 --> Prp18 --> Prp22. All three proteins are released from the spliceosome after step 2 concomitant with release of mature mRNA.

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Elevated levels of Mcm5 in urine sediments are highly predictive of bladder cancer.

OBJECTIVE To investigate whether the expression of P2X3 receptors (implicated in the pathophysiology of pain) is altered in human bladder urothelium from patients with interstitial cystitis (IC, a major symptom of which is pain), and as P2X2 receptors can be co‐expressed with P2X3 receptors, to assess their expression also. PATIENTS AND METHODS Bladder tissue samples were collected from patients undergoing cystectomy or radical prostatectomy. Patients with IC were diagnosed using the international criteria. RNA protein expression levels of both receptors were evaluated using reverse transcription‐polymerase chain reaction (PCR), real‐time quantitative PCR and Western blot analysis. RESULTS P2X2 was expressed in the human urothelium, in a glycosylated form. There was less gene expression of P2X3 in IC urothelium, whereas P2X2 gene expression was unchanged. This contrasted with the protein expression, which was increased for both P2X2 and P2X3. CONCLUSION This is the first report of the expression of the P2X2 receptor in human bladder urothelium. There was greater protein expression of both P2X2 and P2X3 in IC bladder urothelium which did not directly correlate with the gene expression. Changes in expression of P2X2 and P2X3 receptors may contribute to the pain that patients with IC have, and might provide novel drug targets.