Movatterモバイル変換

[0]ホーム
URL:

Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
Thehttps:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

NIH NLM Logo
Log inShow account info
Access keysNCBI HomepageMyNCBI HomepageMain ContentMain Navigation
pubmed logo
Advanced Clipboard
User Guide

Full text links

Nature Publishing Group full text link Nature Publishing Group Free PMC article
Full text links

Actions

Share

.2020 Jun 12;11(1):2977.
doi: 10.1038/s41467-020-16786-5.

A kinase-deficient NTRK2 splice variant predominates in glioma and amplifies several oncogenic signaling pathways

Affiliations

A kinase-deficient NTRK2 splice variant predominates in glioma and amplifies several oncogenic signaling pathways

Siobhan S Pattwell et al. Nat Commun..

Abstract

Independent scientific achievements have led to the discovery of aberrant splicing patterns in oncogenesis, while more recent advances have uncovered novel gene fusions involving neurotrophic tyrosine receptor kinases (NTRKs) in gliomas. The exploration of NTRK splice variants in normal and neoplastic brain provides an intersection of these two rapidly evolving fields. Tropomyosin receptor kinase B (TrkB), encoded NTRK2, is known for critical roles in neuronal survival, differentiation, molecular properties associated with memory, and exhibits intricate splicing patterns and post-translational modifications. Here, we show a role for a truncated NTRK2 splice variant, TrkB.T1, in human glioma. TrkB.T1 enhances PDGF-driven gliomas in vivo, augments PDGF-induced Akt and STAT3 signaling in vitro, while next generation sequencing broadly implicates TrkB.T1 in the PI3K signaling cascades in a ligand-independent fashion. These TrkB.T1 findings highlight the importance of expanding upon whole gene and gene fusion analyses to include splice variants in basic and translational neuro-oncology research.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1. Gene expression differences in human normal brain and human glioma.
a Principal component analysis (PCA) plot of all brain samples from Genotype-Tissue Expression (GTEx) project. Samples from 13 regions fall into two groups clearly separating the cerebellum samples from the rest of the brain regions.b Principal Component analysis (PCA) plot of pooled brain samples from GTEx (with cerebellum and spinal cord removed) and The Cancer Genome Atlas (TCGA) samples using gene expression from 20,214 genes from hg19. Two clear clusters were formed, TCGA-GBM and TCGA-LGG samples fall into one cluster while the supratentorial regions from GTEx maintain the one pooled normal brain cluster froma.c PCA plot of pooled normal brain regions broken down shows minimal regional specificity and clustering based on 20,214 genes from hg19.d PCA plot showing tumor samples from TCGA using gene expression of  20,214 genes in hg19 colored by CpG Island Methylation Phenotype (CIMP) status. CIMP and non-CIMP status for TCGA samples were obtained from Bolouri et al., and show distinct CIMP vs non-CIMP clusters based on total gene expression. Normal brain regions are displayed as circles, brain tumors are displayed as triangles.
Fig. 2
Fig. 2.NTRK2 splice variant, TrkB.T1 is the predominant TrkB isoform, in human glioma.
a Boxplots for transcript expression of TrkB.FL and TrkB.T1 from brain samples from GTEx project (n = 956) and TCGA (n = 530) LGG and TCGA GBM (n = 166) demonstrate a predominance of TrkB.T1 in glioma samples compared with normal brain. TrkB.T1 transcript displayed in teal, TrkB.FL transcript displayed in orange.b Principal Component analysis (PCA) plot of all brain samples from GTEx project and TCGA samples using gene expression from 20,214 genes from hg19. Samples are colored based on status of predominantNTRK2 transcript: teal for TrkB.T1 and orange for TrkB.FL. Normal brain regions are displayed as circles, brain tumors are displayed as triangles.c TrkB.T1 and TrkB.FL transcript expression shows increased expression of TrkB.T1 compared with TrkB.FL across 50 human glioblastoma stem cell (GSC) lines (6 lines: BTSC349, BTSC349, h543, h516, h561, h676; 44 lines from ref. ) (n = 50 for each isoform (TrkB.T1 and TrkB.FL);t-test,p = 1.9 × 10−08). Fora andc, data are represented as boxplots where the middle line is the median, the lower and upper hinges correspond to the first and third quartiles (the 25th and 75th percentiles), the upper whisker extends from the hinge to the largest value no further than 1.5 * IQR from the hinge (where IQR is the inter-quartile range, or distance between the first and third quartiles) and the lower whisker extends from the hinge to the smallest value at most 1.5 * IQR of the hinge while data beyond the end of the whiskers are outlying points that are plotted individually.
Fig. 3
Fig. 3. GO enrichment analysis forNTRK2 in GBM or LGG.
a Schematic of DGCA analysis for the top 350 genes significantly positively correlated withNTRK2 in LGG or GBM. GO terms for biological processes in top 350 differentially expressed genes in GBM compared with normal brain (b) and GO terms for the top 350 differentially expressed genes in the cellular component in GBM compared with normal brain (c) revealed a predominance of genes implicated in morphogenesis and proliferation, as well endocytic compartments and vesicular transport.d Gene ontology enrichment terms for the genesets within the reactome pathways in top 350 differentially expressed genes in LGG compared with normal brain.
Fig. 4
Fig. 4. Human and rodent TrkB.T1 immunostaining in normal brain and glioma.
a Receptor schematic of TrkB.FL (kinase domain demonstrated in orange with phosphorylation sites in yellow) and TrkB.T1 highlighting region of antibody specificity (11-amino acid region shown in teal blue). Extracellular domain, transmembrane domain, and first intracellular amino acids common to both variants shown in light green. TrkB.T1 immunohistochemistry shows punctate vesicular staining in normal human (b) and mouse cortex (c) compared with intense diffuse staining in human glioma (b) and mouse glioma (c) with lack of punctate vesicular pattern. Lack of staining in rodent negative control (no primary antibody) and TrkB.T1−/− cortex demonstrates antibody specificity for TrkB.T1 splice variant. (IDHmut—IDH mutant; 1p19q codel—1p 19 q co-deleted). Immunohistochemistry was performed on independent biological samples of each tumor type, in replicates of 3–5. Representative images were chosen and additional images are shown in Supplementary Fig. 4–6. Photomicrographs are as specified at 40×(scale bar = 200 µm), 200×(scale bar = 50 µm), and 600×(scale bar = 20 µm).
Fig. 5
Fig. 5. TrkB.T1 enhances PDGF signaling in vivo and in vitro.
a Kaplan–Meier plot showing symptom-free survival of PDGFB (orange) vs PDGFB + TrkB.T1 (blue) induced gliomas (median survival 34 days vs 109 days; Mantel-Cox/log-rank hazard ratio 2.306, 95% confidence interval 0.8999 to 5.909,p = 0.0452.b Western blot showing representative efficiency of RCAS-TrkB.T1 in 3T3/tv-a cells (lanes 4, 6, 8) in relation to normal mouse brain (lane 1) and TrkB.T1−/− brain control (lane 2).c Representative western blot showing TrkB.T1-expressing 3T3/tv-a cells exhibit enhanced pSTAT3, pAKT, pS6rp, pERK signaling in response to 10 ng/ml PDGF-BB treatment over the course of 36 h; SS (serum starved), with beta-actin loading control; 3T3/tv-a PDGF-BB treatment time-points repeated in duplicate or triplicate.d TrkB.T1-expressing 3T3/tv-a cells show enhanced phosphorylation of PDGFR-β (Y1021) and maintain expression of total PDGFR-β upon ligand stimulation. Western blotting was performed in triplicate on biologically independent samples, in duplicate using technical replicates, and representative images were chosen.
Fig. 6
Fig. 6. DE analysis reveals distinct gene expression patterns between TrkB.T1 High vs TrkB.T1 Low GSC lines.
a Schematic for differential gene expression analysis between TrkB.T1 High (classical/proneural replicates 448 1–3; classical/mesenchymal G14 replicates 1–3) and TrkB.T1 Low (classical/proneural replicates 559 replicates 1–3; classical/mesenchymal G179 replicates 1–3).b PCA plots show distinct clusters for the TrkB.T1 High GSC line (488) compared with the TrkB.T1 Low GSC (559) line in the classical/proneural lines and distinct and (c) for the TrkB.T1 High GSC Line (G14) compared with the TrkB.T1 Low GSC line (559). GO analysis revealed distinct genesets downregulated in the cellular compartment (d) and distinct genesets downregulated in biological processes (e) in the TrkB.T1 High (448 and G14) line compared with the TrkB.T1 Low (559 and G179) lines. TrkB.T1 Low lines (559 and G179) shown in green and TrkB.T1 High lines (448 and G14) shown in purple.
Fig. 7
Fig. 7. TrkB.T1 upregulates genes involved in PI3K/Akt signaling in human NSCs.
a Cluster dendogram and principal component analysis (PCA) plots (b) show that TrkB.T1-transduced, TrkB.FL-transduced, and GFP-transduced NSC lines cluster independently from each other. Heatmap displays differentially expressed genes between TrkB.T1, TrkB.FL, and GFP infected NSCs (c). Reactome terms for genes upregulated in TrkB.T1 infected NSCs compared with TrkB.FL infected NSCs revealed genes involved in PI3K/Akt, PI3K/ERBB2/ERBB4 signaling pathways (d). GFP-transduced NSCs displayed on PCA plot in bright green, TrkB.T1-transduced cells displayed in teal, TrkB.FL-transduced shows displayed in orange.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Dvinge H, Kim E, Abdel-Wahab O, Bradley RK. RNA splicing factors as oncoproteins and tumour suppressors. Nat. Rev. Cancer. 2016;16:413–430. - PMC - PubMed
    1. Singh B, Eyras E. The role of alternative splicing in cancer. Transcription. 2017;8:91–98. - PMC - PubMed
    1. Oltean S, Bates DO. Hallmarks of alternative splicing in cancer. Oncogene. 2014;33:5311–5318. - PubMed
    1. Jones, K. A., Bossler, A. D., Bellizzi, A. M. & Snow, A. N. BCR-NTRK2 fusion in a low-grade glioma with distinctive morphology and unexpected aggressive behavior. Cold Spring Harb. Mol. Case Stud.5, a003855 (2019). - PMC - PubMed
    1. Gatalica Z, Xiu J, Swensen J, Vranic S. Molecular characterization of cancers with NTRK gene fusions. Mod. Pathol. 2019;32:147–153. - PubMed

Publication types

MeSH terms

Substances

Related information

Grants and funding

LinkOut - more resources

Full text links
Nature Publishing Group full text link Nature Publishing Group Free PMC article
Cite
Send To

NCBI Literature Resources

MeSHPMCBookshelfDisclaimer

The PubMed wordmark and PubMed logo are registered trademarks of the U.S. Department of Health and Human Services (HHS). Unauthorized use of these marks is strictly prohibited.

[8]ページ先頭
©2009-2025 Movatter.jp