A multi-faceted approach to unravel coding and non-coding gene fusions and target chimeric proteins in ataxia.

Ataxia represents a heterogeneous group of neurodegenerative disorders characterized by a loss of balance and coordination, often resulting from mutations in genes vital for cerebellar function and maintenance. Recent advances in genomics have identified gene fusion events as critical contributors to various cancers and neurodegenerative diseases. However, their role in ataxia pathogenesis remains largely unexplored. Our study Hdelved into this possibility by analyzing RNA sequencing data from 1443 diverse samples, including cell and mouse models, patient samples, and healthy controls. We identified 7067 novel gene fusions, potentially pivotal in disease onset. These fusions, notably in-frame, could produce chimeric proteins, disrupt gene regulation, or introduce new functions. We observed conservation of specific amino acids at fusion breakpoints and identified potential aggregate formations in fusion proteins, known to contribute to ataxia. Through AI-based protein structure prediction, we identified topological changes in three high-confidence fusion proteins-TEN1-ACOX1, PEX14-NMNAT1, and ITPR1-GRID2-which could potentially alter their functions. Subsequent virtual drug screening identified several molecules and peptides with high-affinity binding to fusion sites. Molecular dynamics simulations confirmed the stability of these protein-ligand complexes at fusion breakpoints. Additionally, we explored the role of non-coding RNA fusions as miRNA sponges. One such fusion, RP11-547P4-FLJ33910, showed strong interaction with hsa-miR-504-5p, potentially acting as its sponge. This interaction correlated with the upregulation of hsa-miR-504-5p target genes, some previously linked to ataxia. In conclusion, our study unveils new aspects of gene fusions in ataxia, suggesting their significant role in pathogenesis and opening avenues for targeted therapeutic interventions.Communicated by Ramaswamy H. Sarma.

Doxorubicin induces prolonged DNA damage signal in cells overexpressing DEK isoform-2.

DEK has a short isoform (DEK isoform-2; DEK2) that lacks amino acid residues between 49-82. The full-length DEK (DEK isoform-1; DEK1) is ubiquitously expressed and plays a role in different cellular processes but whether DEK2 is involved in these processes remains elusive. We stably overexpressed DEK2 in human bone marrow stromal cell line HS-27A, in which endogenous DEKs were intact or suppressed via short hairpin RNA (sh-RNA). We have found that contrary to ectopic DEK1, DEK2 locates in the nucleus and nucleolus, causes persistent γH2AX signal upon doxorubicin treatment, and couldn't functionally compensate for the loss of DEK1. In addition, DEK2 overexpressing cells were more sensitive to doxorubicin than DEK1-cells. Expressions of DEK1 and DEK2 in cell lines and primary tumors exhibit tissue specificity. DEK1 is upregulated in cancers of the colon, liver, and lung compared to normal tissues while both DEK1 and DEK2 are downregulated in subsets of kidney, prostate, and thyroid carcinomas. Interestingly, only DEK2 was downregulated in a subset of breast tumors suggesting that DEK2 can be modulated differently than DEK1 in specific cancers. In summary, our findings show distinct expression patterns and subcellular location and suggest non-overlapping functions between the two DEK isoforms.

DEK protein level is a biomarker of CD138positive normal and malignant plasma cells.

Overexpression of DEK oncogene is associated with increased proliferation of carcinoma cells and it is observed in several solid tumors due to the amplification of the 6p22.3 chromosomal region where DEK locates. Although the same chromosomal amplification occurs in multiple myeloma (MM), a plasma cell neoplasm, whether the expression and the copy number of the DEK gene are affected in MM remains elusive. We show that despite the increased copy number in CD138positive MM cells (4 out of 41 MM samples), DEK mRNA expression was down-regulated compared with that in CD138negative bone marrow (BM) cells of the same patients (P<0.0001). DEK protein was not detectable by immunohistochemistry (IHC) in CD138positive normal plasma cells or in malignant plasma cells of MM patients (n = 56) whereas it was widely expressed in normal and neoplastic B-cells. Stable knockdown or overexpression of DEK in CD138positive MM cell lines did not affect the proliferation and viability of the cells profoundly in the presence or absence of chemotherapeutic agent melphalan whereas knockdown of DEK moderately but significantly increased the expression level of CD138 (p<0.01). Decreased DEK expression in plasma cells suggests a potential role of this gene in plasma cell development and lack of detectable DEK protein by IHC could be used as a biomarker for normal and malignant plasma cells.

High MN1 expression increases the in vitro clonogenic activity of primary mouse B-cells.

The MN1 (Meningioma 1) gene is overexpressed in certain subtypes of acute myeloid leukemia (AML) and high levels of MN1 expression in mouse bone marrow cells results in myeloid leukemia. We showed that compared with control bone marrow (BM) MN1 expression was increased (2-fold or more) in 29 out of 73 (40%) pediatric B-cell acute lymphoblastic leukemia (B-ALL) patient BM. Additional analysis of MN1 expression in sub-groups within our cohort carrying different chromosome translocations showed that carriers of the good prognostic marker t(12;21)(TEL-AML1) (n=27) expressed significantly more MN1 than both healthy controls (n=9) (P=0.02) and the group carrying the t(9;22)(BCR-ABL) (n=9) (P=0.001). In addition, AML1 expression was also upregulated in 31 out of 45 (68%) B-ALL patient BM compared with control and there was a significant correlation between MN1 and AML1 expression (r=0.3552, P=0.0167). Retroviral MN1 overexpression increased the colony forming activity of mouse Pro-B/Pre-B cells in vitro. Our results suggest that deregulated MN1 expression contributes to the pathogenesis of pediatric B-ALL. Further investigation into the clinical and biological significance of elevated MN1 expression in TEL-AML1(positive) leukemia might provide insight into additional molecular mechanisms contributing to B-ALL and may lead to improved treatment options for patients.