Expression of WDR79 is associated with TP53 mutation and poor prognosis in surgically resected non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) represents a major health burden globally. WD repeat protein 79 (WDR79) is a member of the WD-repeat protein family. WDR79 is a highly conserved and natural antisense transcript to TP53 gene and involved in carcinogenesis of various types of cancer. Whether the alterations of WDR79 protein expression are associated with TP53 mutation and clinicopathological and prognostic implications in the patients with surgically resected NSCLC have not been reported. The purposes of the present study are to investigate the association between the expression of WDR79 and mutant p53 (mtp53) and clinicopathological features in NSCLC by immunohistochemistry. The results showed that positive expression of WDR79 (58.8%, 170/289) and mtp53 (48.1%, 139/289) in NSCLC was significantly higher than that in non-cancerous control lung tissues (5.7%, 3/53 and 1.9%, 1/53, respectively). There was a significantly higher positive percentage of WDR79 expression in NSCLC with lymph node metastasis. The statistically positive correlation between WDR79 and mtp53 expression (r = 0.212, P=0.014) was identified by Spearman's rank correlation analysis. Kaplan-Meier survival curve analysis indicated that positive expression of WDR79 and common positive expression of WDR79 and mtp53 were correlated with poor overall survival rates in NSCLC patients (P = 0.029 and P = 0.041, respectively). Multivariate Cox regression analysis further identified that WDR79 positive expression was an independent unfavorable prognostic factor of NSCLC (P = 0.034). Taken together, positive expression of WDR79 proteins may be related with TP53 mutations and act as valuable independent biomarker to predict poor prognosis of patients with surgically resected NSCLC.

WDR79/TCAB1 plays a conserved role in the control of locomotion and ameliorates phenotypic defects in SMA models.

SMN (Survival Motor Neuron) deficiency is the predominant cause of spinal muscular atrophy (SMA), a severe neurodegenerative disorder that can lead to progressive paralysis and death. Although SMN is required in every cell for proper RNA metabolism, the reason why its loss is especially critical in the motor system is still unclear. SMA genetic models have been employed to identify several modifiers that can ameliorate the deficits induced by SMN depletion. Here we focus on WDR79/TCAB1, a protein important for the biogenesis of several RNA species that has been shown to physically interact with SMN in human cells. We show that WDR79 depletion results in locomotion defects in both Drosophila and Caenorhabditis elegans similar to those elicited by SMN depletion. Consistent with this observation, we find that SMN overexpression rescues the WDR79 loss-of-function phenotype in flies. Most importantly, we also found that WDR79 overexpression ameliorates the locomotion defects induced by SMN depletion in both flies and worms. Our results collectively suggest that WDR79 and SMN play evolutionarily conserved cooperative functions in the nervous system and suggest that WDR79/TCAB1 may have the potential to modify SMA pathogenesis.

Phosphorylation of the Cajal body protein WRAP53ß by ATM promotes its involvement in the DNA damage response.

The cellular response to DNA double-strand breaks is orchestrated by the protein kinase ATM, which phosphorylates key actors in the DNA repair network. WRAP53ß is a multifunctional protein that controls trafficking of factors to Cajal bodies, telomeres and DNA double-strand breaks but what regulates the involvement of WRAP53ß in these separate processes remains unclear. Here, we show that in response to various types of DNA damage, including IR and UV, WRAP53ß is phosphorylated on serine residue 64 by ATM with a time-course that parallels its accumulation at DNA lesions. Interestingly, recruitment of phosphorylated WRAP53ß (pWRAP53ßS64) to sites of such DNA damage promotes its interaction with γH2AX at these locations. Moreover, pWRAP53ßS64 stimulates the accumulation of the repair factor 53BP1 at DNA double-strand breaks and enhances repair of this type of damage via homologous recombination and non-homologous end joining. At the same time, phosphorylation of WRAP53ß is dispensable for its localization to Cajal bodies, where it accumulates even in unstressed cells. These findings not only reveal ATM to be an upstream regulator of WRAP53ß, but also indicates that phosphorylation of WRAP53ß at serine 64 controls its involvement in the DNA damage response and may also restrict its other functions.

On the road with WRAP53ß: guardian of Cajal bodies and genome integrity.

The WRAP53 gene encodes both an antisense transcript (WRAP53α) that stabilizes the tumor suppressor p53 and a protein (WRAP53ß) involved in maintenance of Cajal bodies, telomere elongation and DNA repair. WRAP53ß is one of many proteins containing WD40 domains, known to mediate a variety of cellular processes. These proteins lack enzymatic activity, acting instead as platforms for the assembly of large complexes of proteins and RNAs thus facilitating their interactions. WRAP53ß mediates site-specific interactions between Cajal body factors and DNA repair proteins. Moreover, dysfunction of this protein has been linked to premature aging, cancer and neurodegeneration. Here we summarize the current state of knowledge concerning the multifaceted roles of WRAP53ß in intracellular trafficking, formation of the Cajal body, DNA repair and maintenance of genomic integrity and discuss potential crosstalk between these processes.

The proximity ligation assay reveals that at DNA double-strand breaks WRAP53ß associates with γH2AX and controls interactions between RNF8 and MDC1.

We recently demonstrated that WRAP53ß acts as a key regulator of ubiquitin-dependent repair of DNA double-strand breaks. Here, we applied the proximity ligation assay (PLA) to show that at such breaks WRAP53ß accumulates in close proximity to γH2AX and, furthermore as demonstrated by their co-immunoprecipitation (IP) binds to γH2AX, in a manner dependent on the ATM and ATR kinases. Moreover, formation of complexes between MDC1 and both its partners RNF8 and phosphorylated ATM was visualized. The interaction of MDC1 with RNF8, but not with ATM requires WRAP53ß, suggesting that WRAP53ß facilitates the former interaction without altering phosphorylation of MDC1 by ATM. Furthermore, our findings highlight PLA as a more sensitive method for the analysis of recruitment of repair factors and complex formation at DNA breaks that are difficult to detect using conventional immunofluorescence.

Novel small Cajal-body-specific RNAs identified in Drosophila: probing guide RNA function.

The spliceosomal small nuclear RNAs (snRNAs) are modified post-transcriptionally by introduction of pseudouridines and 2'-O-methyl modifications, which are mediated by box H/ACA and box C/D guide RNAs, respectively. Because of their concentration in the nuclear Cajal body (CB), these guide RNAs are known as small CB-specific (sca) RNAs. In the cell, scaRNAs are associated with the WD-repeat protein WDR79. We used coimmunoprecipitation with WDR79 to recover seven new scaRNAs from Drosophila cell lysates. We demonstrated concentration of these new scaRNAs in the CB by in situ hybridization, and we verified experimentally that they can modify their putative target RNAs. Surprisingly, one of the new scaRNAs targets U6 snRNA, whose modification is generally assumed to occur in the nucleolus, not in the CB. Two other scaRNAs have dual guide functions, one for an snRNA and one for 28S rRNA. Again, the modification of 28S rRNA is assumed to take place in the nucleolus. These findings suggest that canonical scaRNAs may have functions in addition to their established role in modifying U1, U2, U4, and U5 snRNAs. We discuss the likelihood that processing by scaRNAs is not limited to the CB.