A novel protein encoded by circHNRNPU promotes multiple myeloma progression by regulating the bone marrow microenvironment and alternative splicing

Abstract

Backgroud Multiple myeloma (MM) is an incurable plasma cell malignancy in the bone marrow (BM), while immunoglobulin D type of MM (IgD MM) is a very rare but most severe subtype in all MM cases. Therefore, systemic study on IgD MM is purposeful to disclose the recurrent and refractory features in both IgD and other types of MM, and beneficial to the development of potent therapeutic strategy on MM. Methods Agilent SBC-ceRNA microarray chips were employed to examine 3 normal plasma cell samples (NPCs), 5 lgD MM samples and 5 lgG MM samples, respectively. Sanger sequencing, RNase R digestion and qPCR assays were used to detect the existence and expression of circHNRNPU. BaseScope™ RNA ISH assay was performed to test circHNRNPU levels in paraffin-embedded MM tissues. The protein encoded by circHNRNPU was identified by LC-MS/MS, which was named as circHNRNPU_603aa. The function of circHNRNPU_603aa on cellular proliferation and cell cycle was assessed by MTT test, colony formation assay, flow cytometry and MM xenograft mouse model in vivo. RIP-seq, RIP-PCR and WB analysis for ubiquitination were performed to explore the potential mechanism of circHNRNPU_603aa in MM. Exosomes were isolated from the culture supernatant of MM cells by ultracentrifugation and characterized by Transmission Electron Microscope and WB confirmation of exosomes markers Alix and CD9. Results CircHNRNPU was one of the top most abundant and differentially expressed circRNA in IgD MM relative to lgG and NPCs samples. Increased circHNRNPU was associated with poor outcomes in four independent MM patient cohorts. Intriguingly, MM cells secreted circHNRNPU, which encoded a protein named as circHNRNPU_603aa. Overexpressed circHNRNPU_603aa promoted MM cell proliferation in vitro and in vivo, in contrast knockdown of circHNRNPU_603aa by siRNA abrogated these effects. Due to circHNRNPU_603aa including RNA-binding RGG-box region, it regulated SKP2 exon skipping, thereby competitively inhibited c-Myc ubiquitin so as to stabilize c-Myc in MM. MM cells secreted circHNRNPU through exosomes to interfere with various cells in the BM microenvironment. Conclusion Our findings demonstrate that circHNRNPU_603aa is a promising diagnostic and therapeutic marker in both MM cells and BM niche.

mRNA, lncRNA, and circRNA expression profiles in a new aortic dissection murine model induced by hypoxia and Ang II

“…Consistently, these features were reported by Humphrey et al ( 24 ). To demonstrate the potential changes in transcript levels, we performed the SBC-ceRNA microarray assay, which is an effective method to study the molecular mechanism of disease ( 25 – 27 ). A total of 1,742 mRNAs, 2,625 lncRNAs, and 537 circRNAs were dysregulated in OSA-AD tissues, including 1,152 downregulated mRNAs, 590 upregulated mRNAs, 1,507 downregulated lncRNAs, 1,118 upregulated lncRNAs, 167 downregulated circRNAs, and 370 upregulated circRNAs.…”

Comprehensive elaboration of circular RNA in multiple myeloma

“…Further evidence confirmed that circ_SEC61A1 sponged miR-660-5p to increase CDK6 expression, promoting the proliferation and metastasis and restraining the apoptosis of MM cells ( Luo et al, 2021 ). Recently, circHNRNPU, encoding a novel protein named as circHNRNPU_603aa, was associated with poor outcomes in four independent MM patient cohorts, and overexpressed circHNRNPU_603aa promoted MM cell proliferation by regulating the bone marrow microenvironment and alternative splicing ( Tang et al, 2022 ). These results collectively indicate that aberrant circRNA expression is involved in MM cell proliferation, migration, invasion and apoptosis, ultimately affecting cell differentiation and progression.…”

Functions and clinical significance of circular RNAs in acute myeloid leukemia

“…CircRNAs could be translated into proteins through internal ribosome entry site (IRES) in a cap-independent manner, m 6 A-dependent initiation of translation and rolling circle translation ( Figure 2D ). For instance, endogenous circRNAs, such as viral circRNA_000048, circ-E-Cad and circHNRNPU, can translate to a micropeptide vsp21, C-E-Cad-254aa and MAPK1-109aa through IRES-dependent manner ( Gao et al, 2021 ; Tang et al, 2022 ; Zhang et al, 2022 ) ( Figure 2D ), respectively. Moreover, circE7 and circARHGAP3 can translate to E7 protein and circARHGAP35-encoded protein by m 6 A-dependent initiation of translation ( Zhao et al, 2019 ; Li Y. et al, 2021 ) ( Figure 2D ), respectively.…”