Rare Case of de Novo 2p15 Microdeletion Syndrome with Deletion Covering XPO1 and USP34 Genes Diagnosed in a Child - A Case Report.

Introduction: 2p15p16.1 microdeletion syndrome was described for the first time in 2007. The size of the microdeletion is variable and encompasses several genes, like XPO1, USP34, BCL11A, REL, PAPOLG, PEX13, COMMD1, B3GNT2, and EHBP1. Features of the syndrome include short stature, microcephaly, hypotonia, psychomotor developmental delay, anomalies of the fingers of the upper and lower limbs, dysmorphic features like receding forehead, broad nasal bridge, telecanthus, ptosis, flat philtrum, small mouth with a high, narrow palate and everted lower lip. The precise genotype-phenotype correlation in 2p15 deletion syndrome is not understood. The aim of the study is to present the patient's medical history and the diagnostic process. Case Presentation: A boy aged 9 was admitted to the Genetic Outpatient Clinic due to dysmorphic features and mild mental retardation. Full lips, broad nasal root, very light hair, excessively developed subcutaneous tissue, joint laxity, postural defect, flat-valgus foot with sandal gap, brachydactyly, and problems with concentration, memory, and counting were found. Diagnosis was based on microarray testing, and copy-number variation analysis was performed using CytoScan 750K array. Genetic imbalance in the form of a deletion within the short arm of chromosome 2 in the 2p15 region (containing 50 kbp) was shown. The microdeletion covers 2 genes: USP34 and XPO1. Parents were not carriers of that mutation. Conclusion: The phenotypic features presented by the patient were reflected in the genetic test. 2p15 microdeletion syndrome is genetically heterogeneous with possible de novo occurrence, as in the presented case. The precise genotype-phenotype correlation in 2p15 deletion syndrome should be widely studied because in the literature there is mainly mentioned 2p15p16.1 syndrome. Even though 2p15 microdeletion syndrome is a rare discovery and its features are mainly mild, it is necessary to pay special attention to them to refer patients to genetic counseling to make an accurate diagnosis.

Genomic signatures and prognosis of advanced stage Chinese pediatric T cell lymphoblastic lymphoma by whole exome sequencing.

Objective: To investigate the genomic signatures and prognosis of advanced-stage T cell lymphoblastic lymphoma (T-LBL) and to examine the relationship between T-LBL and T cell acute lymphoblastic leukemia (T-ALL). Methods: 35 Chinese T-LBL children with stage III or IV disease were recruited for this study. They were treated with combination chemotherapy and whole exome sequencing. The relationship of the clinical features, prognosis and specific gene mutations was researched. Gene chips of T-LBL and T-ALL were downloaded from a database, and differential gene expression was analyzed. Results: Germline causal gene mutations (CARS or MAP2K2) were detected in 2 patients; 3.06 ± 2.21 somatic causal gene mutations were identified in the 35 patients, and somatic mutations were observed in the NOTCH1, FBXW7, PHF6 and JAK3 genes. NOTCH1 mutations were significantly associated with FBXW7 mutations, and the age at diagnosis of patients with NOTCH1-FBXW7 mutations was less than that of patients without such mutations (P < 0.05). 32 patients achieved complete remission (CR), and 14 and 18 patients were classified into the intermediate risk (IR) group and high risk (HR) group. During a median follow-up of 44 months, 3 patients relapsed. Three-year prospective event free survival (pEFS) was 82.286%, and no significant differences of pEFS were found for different sexes, ages, or statuses of NOTCH1-FBXW7 mutations, (P > 0.05); however, the mean survival time of the IR group was longer than that of the HR group (P < 0.05). Differential expression of genes in the T-LBL and/or T-ALL datasets was analyzed using the R package limma, and 1/3 of the differentially expressed genes were found in both the T-ALL and T-LBL datasets. High expression of PI3K-Akt signal pathway genes and the USP34 gene was found in the T-LBL dataset. Conclusion: Although T-ALL and T-LBL both originate from precursor T-cells and are considered different manifestations of the same disease and the outcome of T-LBL is favorable when using T-ALL-based chemotherapy, there are differences in the gene distribution between T-LBL and T-ALL. It seems that the PI3K-Akt signaling pathway and the USP34 gene play important roles in T-LBL, but medicines targeting the USP34 gene or the PI3K-Akt pathway may be invalid.

Ubiquitin-specific protease 34 in macrophages limits CD8 T cell-mediated onset of vitiligo in mice.

As an autoimmune disorder, vitiligo is characterized by depigmented skin macules. CD8+T cells and macrophages enrichment promote the onset of vitiligo, while the role of macrophages to CD8+T is not well deciphered. To develop a mouse model of vitiligo with prominent epidermal depigmentation, Krt14-Kitl* transgenic mice containing an elevated number of melanocytes in the epidermis with membrane-bound Kit ligand (Kitl*) were adoptively transferred with premelanosome protein (PMEL) CD8+ T cells. On the other hand, Krt14-Kitl* mice were mated with ubiquitin-specific protease 34 (USP34)MKO mice to decipher the role of USP34 in vitiligo. Vitiligo scores and PMEL CD8+ T cell enrichment were detected with flow cytometry. Human peripheral blood mononuclear cells (PBMCs) or mice bone marrow-derived macrophages (BMDMs) were incubated with lipopolysaccharide (LPS), CpG, or co-incubated with KU-55933, an ataxia telangiectasia-mutated (ATM) inhibitor. Chemokine (C-C motif) ligand 2 (CCL2), Ccl5, and interleukin (Il)-12α expression was assayed with real-time PCR, and p-IKKα/ß was assayed with Western blots. USP34 was up-regulated in the PBMCs of vitiligo patients and LPS-stimulated BMDMs. USP34 deficiency did not affect the differentiation of CD11b+F4/80+ macrophages in the bone marrow. Immunoprecipitation demonstrated the interaction between USP34 and ATM. USP34 deficiency or KU-55933 administration promoted the induction of Ccl2, Ccl5, Il12α, and p-IKKα/ß in LPS or CpG stimulated BMDMs; KU-55933 administration could not affect the expression of the above molecules in USP34 deficient BMDMs. It further revealed that USP34 deficiency promoted the development of vitiligo with increased PMEL CD8+ T cell enrichment, which was not affected by KU-55933 administration. USP34 deficiency in macrophages promotes the onset of vitiligo with increased PMEL CD8+ T cell enrichment, and USP34/ATM complex can be considered as a therapy target.

Whole-Exome Sequencing Implicates the USP34 rs777591A > G Intron Variant in Chronic Obstructive Pulmonary Disease in a Kashi Cohort.

Genetic factors are important factors in chronic obstructive pulmonary disease (COPD) onset. Plenty of risk and new causative genes for COPD have been identified in patients of the Chinese Han population. In contrast, we know considerably little concerning the genetics in the Kashi COPD population (Uyghur). This study aims at clarifying the genetic maps regarding COPD susceptibility in Kashi (China). Whole-exome sequencing (WES) was used to analyze three Uyghur families with COPD in Kashi (eight patients and one healthy control). Sanger sequencing was also used to verify the WES results in 541 unrelated Uyghur COPD patients and 534 Uyghur healthy controls. WES showed 72 single nucleotide variants (SNVs), two deletions, and small insertions (InDels), 26 copy number variants (CNVs), and 34 structural variants (SVs), including g.71230620T > A (rs12449210T > A, NC_000,016.10) in the HYDIN axonemal central pair apparatus protein (HYDIN) gene and g.61190482A > G (rs777591A > G, NC_000002.12) in the ubiquitin-specific protease 34 (USP34) gene. After Sanger sequencing, we found that rs777591"AA" under different genetic models except for the dominant model (adjusted OR = 0.8559, 95%CI 0.6568-1.115, p > .05), could significantly reduce COPD risk, but rs12449210T > A was not related to COPD. In stratified analysis of smoking status, rs777591"AA" reduced COPD risk significantly among the nonsmoker group. Protein and mRNA expression of USP34 in cigarette smoke extract-treated BEAS-2b cells increased significantly compared with those in the control group. Our findings associate the USP34 rs777591"AA" genotype as a protector factor in COPD.

The deubiquitinase USP34 stabilizes SOX2 and induces cell survival and drug resistance in laryngeal squamous cell carcinoma.

Recent studies showed that the deubiquitinase ubiquitin-specific protease 34 (USP34) was involved in the tumorigenesis of several tumors, but its function and mechanism are still unclear in laryngeal squamous cell carcinoma (LSCC). In this study, we found that USP34 and SOX2 were elevated in LSCC tumor tissues, and we also found that USP34 expression was positively correlated with SOX2 expression. Our further studies showed that USP34 regulated the protein level of SOX2 in LSCC cells, but not the mRNA level, which suggested that USP34 stabilized SOX2. Moreover, USP34, as a deubiquitinase, could interact with SOX2, and reduce the polyubiquitination of SOX2. In addition, knockdown of USP34 could significantly inhibit LSCC cell growth, but overexpression of SOX2 could reverse this effect. Finally, we also found that USP34 and SOX2 were upregulated in cisplatin-resistant LSCC cells, but knockdown of USP34 could enhance the drug sensitivity of cisplatin in the resistant cells. Collectively, targeting USP34/SOX2 axis may be a promising strategy for the treatment of LSCC.

Downregulation of USP34 Inhibits the Growth and Migration of Pancreatic Cancer Cells via Inhibiting the PRR11.

BACKGROUND: Pancreatic cancer (PC) is a highly lethal malignancy worldwide. Our previous study indicated that overexpression of USP34 could promote tumor growth in PC cells. Therefore, this study aimed to further investigate the role of USP34 during the tumorigenesis of PC. METHODS: The level of USP34 in PANC-1 and MiaPaCa-2 cells transfected with USP34-shRNAs was detected by RT-qPCR. Moreover, transwell migration and Annexin V/PI analysis were conducted to detect cell migration and apoptosis, respectively. RESULTS: In this study, downregulation of USP34 markedly inhibited proliferation and migration, and induced apoptosis in PANC-1 cells. Moreover, silencing of USP34 obviously downregulated the levels of PRR11 and p-p38 in PANC-1 cells. An in vivo study in nude mice bearing PANC-1 cell xenografts confirmed these results. CONCLUSION: Downregulation of USP34 could inhibit proliferation and migration in PANC-1 cells via inhibiting PRR11, and inactivating p38 MAPK signaling. Therefore, USP34 might be a potential therapeutic target for the treatment of PC.

Systematic analysis of the frequently amplified 2p15-p16.1 locus reveals PAPOLG as a potential proto-oncogene in follicular and transformed follicular lymphoma.

Transformed follicular lymphoma (tFL) originates from histological transformation of follicular lymphoma (FL), which is the most common indolent non-Hodgkin lymphoma. High-resolution genomic copy-number analysis previously identified frequent amplification of the 2p15-p16.1 locus in FL and tFL cases. The genes (i.e. BCL11A, PAPOLG, PUS10, and USP34) in this amplified locus have not been systematically investigated to date in terms of their role in FL pathogenesis or transformation to tFL. Here we investigated the relationship between amplification and expression of genes in 2p15-p16.1 as well as their expression after histological transformation. NCBI GEO SNP array and gene expression profile (GEP) data of tFL cases were analyzed to evaluate the relationship between amplification and mRNA expression. Moreover, transcript levels of these four genes in FL cases were compared with those of patient-matched tFL cases and normal B-cells. Amplification of the 2p15-p16.1 locus is associated with increased transcription of BCL11A and PAPOLG in tFL cases, of which the latter showed increased expression after histological transformation. Compared with the level in normal B-cells, PAPOLG was significantly overexpressed in FL cases, but expression levels of the other three genes did not show any significant difference. Altogether these results suggest that PAPOLG may be the most critical gene in terms of transformation to tFL.

USP34 Regulated Human Pancreatic Cancer Cell Survival via AKT and PKC Pathways.

Pancreatic cancer is known to be a fatal disease, which is difficult to be diagnosed in its early stages. Ubiquitin-Specific Protease 34 (USP34) are closely related to human cancers in the development and progression. However, there are rarely studies about the role of USP34 in pancreatic cancer. Thus, we aimed to investigate the effect of USP34 in human pancreatic cancer. Short-hairpin RNA targeting USP34 (USP34-shRNA) and USP34 overexpression lentivirus were used in the current study. The level of USP34 in human pancreatic cancer (PANC-1) cells were then analyzed by quantitative (q)RT-PCR. In addition, Western blotting was used to examine phosphorylated (p)-AKT, p-protein kinase C (PKC) and p-extracellular signal-regulated kinase (ERK) protein levels. CCK-8 assay, flow cytometry, and migration assay were used to detect cell proliferation, apoptosis and migration, respectively in vitro. According to the result of qRT-PCR and Western blotting, USP34-shRNA1 significantly downregulated USP34 gene level in PANC-1 cell. Subsequently, Western blotting assay indicated that USP34 silencing significantly down-regulated the expression of p-AKT and p-PKC in cells. On the other hand, USP34 overexpressing remarkably up-regulated the expression of p-AKT and p-PKC in cells. In addition, USP34 overexpression promoted PANC-1 cell proliferation and migration via up-regulating the proteins of p-AKT and p-PKC. Moreover, USP34 overexpression reversed AKT inhibitor and PKC inhibitor induced PACN-1 cell apoptosis. Our results indicated USP34 regulated h PANC-1 cell survival via AKT and PKC pathways, and which played a pro-survival role in human pancreatic cancer. Therefore, we suggested USP34 could be a potential therapeutic target for pancreatic cancer.

The ubiquitin specific protease USP34 protects the ubiquitin ligase gp78 from proteasomal degradation.

The E3 ubiquitin (Ub) ligase gp78 plays an important role in endoplasmic reticulum (ER)-associated degradation (ERAD) and regulation of lipid biogenesis. Although a variety of substrates of gp78 have been described, the regulation of the degradation of gp78 itself remains poorly understood. To address this problem, we used co-immunoprecipitation-coupled liquid chromatography-tandem mass spectrometry (Co-IP/LC-MS/MS) to identify novel proteins interacting with gp78. One of the proteins identified in this study is the deubiquitylating (DUB) enzyme USP34 (Ub-specific protease 34). We demonstrate that knockdown of USP34 facilitates proteasomal degradation of gp78 and consequently impairs the function of gp78 in regulating lipid droplet formation. This study unveils a previously unknown function of USP34 in regulating the metabolic stability of gp78 and adds to our understanding of the relevance of partnering of DUBs and E3s in regulation of protein ubiquitylation.

Inhibition of ubiquitin-specific protease 34 (USP34) induces epithelial-mesenchymal transition and promotes stemness in mammary epithelial cells.

Ubiquitin-specific protease 34 (USP34) is a deubiquitinating enzyme that regulates Axin stability and plays a critical role in Wnt/ß-catenin signaling. We sought to investigate the role of USP34 on epithelial-mesenchymal (EMT) induction and its effects on mammary epithelial stem cells. USP34 expression levels were relatively lower in MDA-MB-231 and 4T1 mesenchymal-like cells when compared to epithelial-like cells. Inhibition of USP34 in NMuMG cells induced EMT, as evidenced by the upregulation of EMT markers including N-cadherin, phospho-Smad3, Snail and active-ß-catenin, as well as the downregulation of Axin 1 and E-cadherin. USP34 knockdown (KD) in these cells also resulted in the acquisition of invasive behavior, and promoted stemness as indicated by enhanced mammosphere-forming ability, concomitant with the upregulation of Nanog, Oct4 and Sox2 mRNA expression. Endogenous USP34 expression was observed to be at low levels in virgin mouse mammary glands in vivo. When USP34-KD cells were transplanted into the cleared mammary fat pads (CFP) of mice, these cells reconstituted the mammary gland with ductal tree development within 3months. Our findings suggest a previously unknown role for USP34 in mammary gland development.

Characteristics of 2p15-p16.1 microdeletion syndrome: Review and description of two additional patients.

Many new microdeletion syndromes have been characterized in the past decade, including 2p15-p16.1 microdeletion syndrome. More than 10 patients with this syndrome have been described. Recently, we encountered two additional patients with 2p15-p16.1 microdeletion syndrome. All patients showed variable degrees of intellectual disability, with the autistic features characteristic of this syndrome. Seven out of 16 patients (44%) showed structural abnormalities in the brain, which is also an important feature of this syndrome. The shortest region of microdeletion overlap among the patients includes two genes, USP34 and XPO1. Although these genes have some functional relevance to cancer, they have not been associated with neurological functions. Diagnosis of additional patients with 2p15-p16.1 microdeletion syndrome and identification of pathogenic mutations in this region will help identify the genes responsible for the neurological features of the syndrome.

Haploinsufficiency of XPO1 and USP34 by a de novo 230 kb deletion in 2p15, in a patient with mild intellectual disability and cranio-facial dysmorphisms.

2p15p16.1-deletion syndrome was first described in 2007 based on the clinical presentation of two patients. The syndrome is characterized by intellectual disability, autism spectrum disorders, microcephaly, dysmorphic facial features and a variety of congenital organ defects. The precise genotype-phenotype correlation in 2p15-deletion syndrome is not understood. However, greater insight can be obtained by thorough clinical investigation of patients carrying deletions, especially those of small size. We report a 21-year-old male patient with features overlapping the clinical spectrum of the 2p15p16.1-deletion syndrome, such as intellectual disability, dysmorphic facial features, and congenital defects. He carried a 230 kb de novo deletion (chr2:61500346-61733075 bp, hg19), which affects the genes USP34, SNORA70B and XPO1. While there is a lack of functional data on SNORA70B, the involvement of USP34 and XPO1 in the regulation of fundamental developmental processes is well known. We suggest that haploinsufficiency of one or both of these genes is likely to be responsible for the disease in our patient.