• BMB Reports
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• 제55권7호
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• pp.348-353
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• 2022

Gastrointestinal cancer is associated with a high mortality rate. Here, we report that the splice variant of NRP/B contributes to tumorigenic activity in highly malignant gastric cancer through dissociation from the tumor repressor, HDAC5. NRP/B mRNA expression is significantly higher in the human gastric cancer tissues than in the normal tissues. Further, high levels of both the NRP/B splice variant and Lgr5, but not the full-length protein, are found in highly tumorigenic gastric tumor cells, but not in non-tumorigenic cells. The loss of NRP/B markedly inhibits cell migration and invasion, which reduces tumor formation in vivo. Importantly, the inhibition of alternative splicing increases the levels of NRP/B-1 mRNA and protein in AGS cells. The ectopic expression of full-length NRP/B exhibits tumor-suppressive activity, whereas NRP/B-2 induces the noninvasive human gastric cancer cells tumorigenesis. The splice variant NRP/B-2 which loses the capacity to interact with tumor repressors promoted oncogenic activity, suggesting that the BTB/POZ domain in the N-terminus has a crucial role in the suppression of gastric cancer. Therefore, the regulation of alternative splicing of the NRP/B gene is a potential novel target for the treatment of gastrointestinal cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제15권5호
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• pp.2153-2158
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• 2014

Beclin 1 is a key factor for initiation and regulation of autophagy, which is a cellular catabolic process involved in tumorigenesis. To investigate the role of alternative splicing of Beclin1 in the regulation of autophagy in leukemia cells, Beclin1 mRNA from 6 different types of cell lines and peripheral blood mononuclear cells from 2 healthy volunteers was reversely transcribed, subcloned, and screened for alternative splicing. New transcript variants were analyzed by DNA sequencing. A transcript variant of Beclin 1 gene carrying a deletion of exon 11, which encoded a C-terminal truncation of Beclin 1 isoform, was found. The alternative isoform was assessed by bioinformatics, immunoblotting and subcellular localization. The results showed that this variable transcript is generated by alternative 3' splicing, and its translational product displayed a reduced activity in induction of autophagy by starvation, indicating that the spliced isoform might function as a dominant negative modulator of autophagy. Our findings suggest that the alternative splicing of Beclin 1 might play important roles in leukemogenesis regulated by autophagy.

• BMB Reports
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• 제41권6호
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• pp.415-434
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• 2008

Phosphoinositide-specific phospholipase C is an effector molecule in the signal transduction process. It generates two second messengers, inositol-1,4,5-trisphosphate and diacylglycerol from phosphatidylinositol 4,5-bisphosphate. Currently, thirteen mammal PLC isozymes have been identified, and they are divided into six groups: PLC-$\beta$, -$\gamma$, -$\delta$, -$\varepsilon$, -$\zeta$ and -$\eta$. Sequence analysis studies demonstrated that each isozyme has more than one alternative splicing variant. PLC isozymes contain the X and Y domains that are responsible for catalytic activity. Several other domains including the PH domain, the C2 domain and EF hand motifs are involved in various biological functions of PLC isozymes as signaling proteins. The distribution of PLC isozymes is tissue and organ specific. Recent studies on isolated cells and knockout mice depleted of PLC isozymes have revealed their distinct phenotypes. Given the specificity in distribution and cellular localization, it is clear that each PLC isozyme bears a unique function in the modulation of physiological responses. In this review, we discuss the structural organization, enzymatic properties and molecular diversity of PLC splicing variants and study functional and physiological roles of each isozyme.

• Molecules and Cells
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• 제41권8호
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• pp.733-741
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• 2018

Mutations in spliceosome components have been implicated in carcinogenesis of various types of cancer. One of the most frequently found is U2AF1 S34F missense mutation. Functional analyses of this mutation have been largely limited to hematological malignancies although the mutation is also frequently seen in other cancer types including lung adenocarcinoma (LUAD). We examined the impact of knockdown (KD) of wild type (wt) U2AF1 and ectopic expression of two splice variant S34F mutant proteins in terms of alternative splicing (AS) pattern and cell cycle progression in A549 lung cancer cells. We demonstrate that induction of distinct AS events and disruption of mitosis at distinct sub-stages result from KD and ectopic expression of the mutant proteins. Importantly, when compared with the splicing pattern seen in LUAD patients with U2AF1 S34F mutation, ectopic expression of S34F mutants but not KD was shown to result in common AS events in several genes involved in cell cycle progression. Our study thus points to an active role of U2AF1 S34F mutant protein in inducing cell cycle dysregulation and mitotic stress. In addition, alternatively spliced genes which we describe here may represent novel potential markers of lung cancer development.

• BMB Reports
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• 제45권9호
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• pp.515-520
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• 2012

High expression of osmotically responsive genes1 (HOS1), a key regulator of low temperature response and flowering time, encodes an E3 ubiquitin ligase in Arabidopsis. Here, we report characterization of a newly identified splice variant (HOS1-L) of HOS1. Comparative analyses revealed that HOS1-L has a longer 5' nucleotide sequence than that of the previously identified HOS1 (HOS1-S) and that its protein sequence was more conserved than that of HOS1-S in plants. HOS1-L transcripts were spatio-temporally more abundant than those of HOS1-S. The recovery rate of HOS1-S expression was faster than that of HOS1-L after cold treatment. Diurnal oscillation patterns of HOS1-L revealed that HOS1-L expression was affected by photoperiod. An in vitro pull-down assay revealed that the HOS1-L protein interacted with the ICE1 protein. HOS1-L overexpression caused delayed flowering in wild-type plants. Collectively, these results suggest regulation of HOS1 expression at the post-transcriptional level.

• 통합자연과학논문집
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• 제6권4호
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• pp.234-243
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• 2013

N-terminal kinase-like (NTKL) protein was initially identified as a protein binding to protein kinase B (PKB, also known as Akt). Though NTKL-BP1 (NTKL-binding protein 1) has been identified as an NTKL binding protein, its functions related to binding have not yet been elucidated. Here, a new alternative spliced variant of NTKL and its association with integrin ${\beta}1$ is described, in addition to the kinase activity of NTKL and its substrate candidates. Although the phosphorylation of the candidates must be further confirmed using other experimental methods, the observation that NTKL can phosphorylate ROCK1, DYRK3, and MST1 indicates that NTKL may act as a signaling protein to regulate actin assembly, cell migration, cell growth, and to facilitate differentiation and development in an integrin-associated manner.

• Molecules and Cells
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• 제21권1호
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• pp.21-28
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• 2006

The interferon-induced, double-stranded RNA (dsRNA)-dependent protein kinase PKR plays a key role in interferon-mediated host defense against viral infection, and is implicated in cellular transformation and apoptosis. We have isolated a cDNA of simian PKR encoding a product with 83% amino acid identity to the human homolog and showed that PKR expression is significantly attenuated in the Vero E6 African green monkey kidney cells devoid of type I interferon genes. A variant form of PKR lacking the exon 12 in the kinase domain is produced in these cells, presumably from an alternatively spliced transcript. Unlike wild type PKR, the variant protein named PKR-${\Delta}E12$ is incapable of auto-phosphorylation and phosphorylation of eIF2-${\alpha}$, indicating that the kinase sub-domains III and IV embedded in exon 12 are indispensable for catalytic function. PKR-${\Delta}E12$ had no dominant negative effect but was weakly phosphorylated in trans by wild type PKR.

• Molecules and Cells
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• 제27권4호
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• pp.467-473
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• 2009

Our previous study suggested that OsBWMK1, a gene which encodes a member of the rice MAP kinase family, generates transcript variants which show distinct expression patterns in response to environmental stresses. The transcript variants are generated by alternative splicing and by use of alternative promoters. To test whether the two alternative promoters, pOsBWMK1L (promoter for the OsBWMK1L splice variant) and pOsBWMK1S (promoter for the OsBWMK1S splice variant), are biologically functional, we analyzed transgenic plants expressing GUS fusion constructs for each promoter. Both pOsBWMK1L and pOsBWMK1S are biologically active, although the activity of pOsBWMK1S is lower than that of pOsBWMK1L. Histochemical analysis revealed that pOsBWMK1L is constitutively active in most tissues at various developmental stages in rice and Arabidopsis, whereas pOsBWMK1S activity is spatially and temporally restricted. Furthermore, the expression of pOsBWMK1S::GUS was upregulated in response to hydrogen peroxide, a plant defense signaling molecule, in both plant species. These results suggest that the differential expression of OsBWMK1 splice variants is the result of alternative promoter usage and, moreover, that the mechanisms controlling OsBWMK1 gene expression are conserved in both monocot and dicot plants.

• Asian-Australasian Journal of Animal Sciences
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• 제31권10호
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• pp.1581-1590
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• 2018

Objective: The aim of this study was to clone alternative splicing isoforms of pig myoneurin (MYNN), predict the structure and function of coding protein, and study temporal and spatial expression characteristics of each transcript. Methods: Alternative splice isoforms of MYNN were identified using RNA sequencing (RNA-seq) and cloning techniques. Quantitative real-time polymerase chain reaction (qPCR) was employed to detect expression patterns in 11 tissues of Large White (LW) and Mashen (MS) pigs, and to study developmental expression patterns in cerebellum (CE), stomach (ST), and longissimus dorsi (LD). Results: The results showed that MYNN had two alternatively spliced isoforms, MYNN-1 (GenBank accession number: KY470829) and MYNN-2 (GenBank accession number: KY670835). MYNN-1 coding sequence (CDS) is composed of 1,830 bp encoding 609 AA, whereas MYNN-2 CDS is composed of 1,746 bp encoding 581 AA. MYNN-2 was 84 bp less than MYNN-1 and lacked the sixth exon. MYNN-2 was found to have one $C_2H_2$ type zinc finger protein domain less than MYNN-1. Two variants were ubiquitously expressed in all pig tissues, and there were significant differences in expression of different tissues (p<0.05; p<0.01). The expression of MYNN-1 was significantly higher than that of MYNN-2 in almost tissues (p<0.05; p<0.01), which testified that MYNN-1 is the main variant. The expression of two isoforms decreased gradually with increase of age in ST and CE of MS pig, whereas increased gradually in LW pig. In LD, the expression of two isoforms increased first and then decreased with increase of age in MS pig, and decreased gradually in LW pig. Conclusion: Two transcripts of pig MYNN were successfully cloned and MYNN-1 was main variant. MYNN was highly expressed in ST, CE, and LD, and their expression was regular. We speculated that MYNN plays important roles in digestion/absorption and skeletal muscle growth, whereas the specific mechanisms require further elucidation.

• Animal cells and systems
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• 제12권3호
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• pp.137-141
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• 2008

Molecular cloning revealed the three isoforms($Ca_v3.1,\;Ca_v3.2,\;and\;Ca_v3.3$) of the T-type calcium channel subfamily. Expression studies exhibited their distinctive electrophysiological and pharmacological properties, accounting for diverse properties of T-type calcium channel currents previously characterized from isolated cells. However, electrophysiological properties of ion channels have shown to be more diversified by their splice variants. We here searched splice variants of rat $Ca_v3.1$ T-type channel by reverse-transcription-polymerase chain reaction(RT-PCR) to further explore diversity of $Ca_v3.1$. Interestingly, analyses of cloned RT-PCR products displayed that there were at least four splicing variants of rat $Ca_v3.1$ in the loop connecting repeats III and IV. Southern blot analyses indicated that the predominantly detected variant in brain was $Ca_v3.1a$(492 bp), which were rarely detected in most of peripheral tissues. Other two variants($Ca_v3.1c$, 546 bp; $Ca_v3.1d$, 525 bp) were detected in most of the tissues examined. The smallest isoform($Ca_v3.1b$, 471 bp) was rarely detected all the tissues. Electrophysiological characterization of the splicing variants indicated that the splice variants differ in inactivation kinetics and the voltage dependence of activation and inactivation as well.