Top

Published in:

Open Access 01-12-2018 | Research article

Regulation of the oncogenic phenotype by the nuclear body protein ZC3H8

Authors: John A. Schmidt, Keith G. Danielson, Emily R. Duffner, Sara G. Radecki, Gerard T. Walker, Amber Shelton, Tianjiao Wang, Janice E. Knepper

Published in: BMC Cancer | Issue 1/2018

Abstract

Background

The Zc3h8 gene encodes a protein with three zinc finger motifs in the C-terminal region. The protein has been identified as a component of the Little Elongation Complex, involved in transcription of small nuclear RNAs. ZC3H8 is overexpressed in a number of human and mouse breast cancer cell lines, and elevated mRNA levels are associated with a poorer prognosis for women with breast cancer.

Methods

We used RNA silencing to decrease levels of expression in mouse mammary tumor cells and overexpression of ZC3H8 in cells derived from the normal mouse mammary gland. We measured characteristics of cell behavior in vitro, including proliferation, migration, invasion, growth in soft agar, and spheroid growth. We assessed the ability of these cells to form tumors in syngeneic BALB/c mice. ZC3H8 protein was visualized in cells using confocal microscopy.

Results

Tumor cells with lower ZC3H8 expression exhibited decreased proliferation rates, slower migration, reduced ability to invade through a basement membrane, and decreased anchorage independent growth in vitro. Cells with lower ZC3H8 levels formed fewer and smaller tumors in animals. Overexpression of ZC3H8 in non-tumorigenic COMMA-D cells led to an opposite effect. ZC3H8 protein localized to both PML bodies and Cajal bodies within the nucleus. ZC3H8 has a casein kinase 2 (CK2) phosphorylation site near the N-terminus, and a CK2 inhibitor caused the numerous PML bodies and ZC3H8 to coalesce to a few larger bodies. Removal of the inhibitor restored PML bodies to their original state. A mutant ZC3H8 lacking the predicted CK2 phosphorylation site showed localization and numbers of ZC3H8/PML bodies similar to wild type. In contrast, a mutant constructed with a glutamic acid in place of the phosphorylatable threonine showed dramatically increased numbers of smaller nuclear foci.

Conclusions

These experiments demonstrate that Zc3h8 expression contributes to aggressive tumor cell behavior in vitro and in vivo. Our studies show that ZC3H8 integrity is key to maintenance of PML bodies. The work provides a link between the Little Elongation Complex, PML bodies, and the cancer cell phenotype.

Available only for authorised users

Dahm K, Nielsen PJ, Muller AM. Transcripts of Fliz1, a nuclear zinc finger protein, are expressed in discrete foci of the murine fetal liver. Genomics. 2001;73(2):194–202.CrossRefPubMed

Gao J, Aksoy BA, Dogrusoz U, Dresdner G, Gross B, Sumer SO, Sun Y, Jacobsen A, Sinha R, Larsson E, et al. Integrative analysis of complex cancer genomics and clinical profiles using the cBioPortal. Sci Signal. 2013;6(269):pl1.CrossRefPubMedPubMedCentral

Zhang M, Han G, Wang C, Cheng K, Li R, Liu H, Wei X, Ye M, Zou H. A bead-based approach for large-scale identification of in vitro kinase substrates. Proteomics. 2011;11(24):4632–7.CrossRefPubMed

Lai WS, Carballo E, Thorn JM, Kennington EA, Blackshear PJ. Interactions of CCCH zinc finger proteins with mRNA. Binding of tristetraprolin-related zinc finger proteins to au-rich elements and destabilization of mRNA. J Biol Chem. 2000;275(23):17827–37.CrossRefPubMed

Kelly SM, Leung SW, Apponi LH, Bramley AM, Tran EJ, Chekanova JA, Wente SR, Corbett AH. Recognition of polyadenosine RNA by the zinc finger domain of nuclear poly(a) RNA-binding protein 2 (Nab2) is required for correct mRNA 3′-end formation. J Biol Chem. 2010;285(34):26022–32.CrossRefPubMedPubMedCentral

Blackshear PJ, Phillips RS, Ghosh S, Ramos SB, Richfield EK, Lai WS. Zfp36l3, a rodent X chromosome gene encoding a placenta-specific member of the Tristetraprolin family of CCCH tandem zinc finger proteins. Biol Reprod. 2005;73(2):297–307.CrossRefPubMed

De J, Lai WS, Thorn JM, Goldsworthy SM, Liu X, Blackwell TK, Blackshear PJ. Identification of four CCCH zinc finger proteins in Xenopus, including a novel vertebrate protein with four zinc fingers and severely restricted expression. Gene. 1999;228(1–2):133–45.CrossRefPubMed

Li J, Jia D, Chen X. HUA1, a regulator of stamen and carpel identities in Arabidopsis, codes for a nuclear RNA binding protein. Plant Cell. 2001;13(10):2269–81.CrossRefPubMedPubMedCentral

Ling AS, Trotter JR, Hendriks EF. A zinc finger protein, TbZC3H20, stabilizes two developmentally regulated mRNAs in trypanosomes. J Biol Chem. 2011;286(23):20152–62.CrossRefPubMedPubMedCentral

10.

Wells ML, Huang W, Li L, Gerrish KE, Fargo DC, Ozsolak F, Blackshear PJ. Posttranscriptional regulation of cell-cell interaction protein-encoding transcripts by Zfs1p in Schizosaccharomyces pombe. Mol Cell Biol. 2012;32(20):4206–14.CrossRefPubMedPubMedCentral

11.

Droll D, Minia I, Fadda A, Singh A, Stewart M, Queiroz R, Clayton C. Post-transcriptional regulation of the trypanosome heat shock response by a zinc finger protein. PLoS Pathog. 2013;9(4):e1003286.CrossRefPubMedPubMedCentral

12.

Baltz AG, Munschauer M, Schwanhausser B, Vasile A, Murakawa Y, Schueler M, Youngs N, Penfold-Brown D, Drew K, Milek M, et al. The mRNA-bound proteome and its global occupancy profile on protein-coding transcripts. Mol Cell. 2012;46(5):674–90.CrossRefPubMed

13.

Wells ML, Hicks SN, Perera L, Blackshear PJ. Functional equivalence of an evolutionarily conserved RNA binding module. J Biol Chem. 2015;290(40):24413–23.CrossRefPubMedPubMedCentral

14.

Hu D, Smith ER, Garruss AS, Mohaghegh N, Varberg JM, Lin C, Jackson J, Gao X, Saraf A, Florens L, et al. The little elongation complex functions at initiation and elongation phases of snRNA gene transcription. Mol Cell. 2013;51(4):493–505.CrossRefPubMedPubMedCentral

15.

Egloff S, Vitali P, Tellier M, Raffel R, Murphy S, Kiss T. The 7SK snRNP associates with the little elongation complex to promote snRNA gene expression. EMBO J. 2017;36(7):934–48.CrossRefPubMedPubMedCentral

16.

Takahashi H, Takigawa I, Watanabe M, Anwar D, Shibata M, Tomomori-Sato C, Sato S, Ranjan A, Seidel CW, Tsukiyama T, et al. MED26 regulates the transcription of snRNA genes through the recruitment of little elongation complex. Nat Commun. 2015;6:5941.CrossRefPubMedPubMedCentral

17.

Fong KW, Li Y, Wang W, Ma W, Li K, Qi RZ, Liu D, Songyang Z, Chen J. Whole-genome screening identifies proteins localized to distinct nuclear bodies. J Cell Biol. 2013;203(1):149–64.CrossRefPubMedPubMedCentral

18.

Singh DK, Prasanth KV. Functional insights into the role of nuclear-retained long noncoding RNAs in gene expression control in mammalian cells. Chromosome res. 2013;21(6–7):695–711.CrossRefPubMedPubMedCentral

19.

Prasanth KV, Prasanth SG, Xuan Z, Hearn S, Freier SM, Bennett CF, Zhang MQ, Spector DL. Regulating gene expression through RNA nuclear retention. Cell. 2005;123(2):249–63.CrossRefPubMed

20.

Kang JJ, Schwegel T, Knepper JE. Sequence similarity between the long terminal repeat coding regions of mammary-tumorigenic BALB/cV and renal-tumorigenic C3H-K strains of mouse mammary tumor virus. Virology. 1993;196(1):303–8.CrossRefPubMed

21.

Slagle BL, Lanford RE, Medina D, Butel JS. Expression of mammary tumor virus proteins in preneoplastic outgrowth lines and mammary tumors of BALB/cV mice. Cancer Res. 1984;44(5):2155–62.PubMed

22.

Knepper JE, Medina D, Butel JS. Activation of endogenous MMTV proviruses in murine mammary cancer induced by chemical carcinogen. Int J Cancer. 1987;40(3):414–22.CrossRefPubMed

23.

Danielson KG, Oborn CJ, Durban EM, Butel JS, Medina D. Epithelial mouse mammary cell line exhibiting normal morphogenesis in vivo and functional differentiation in vitro. Proc Natl Acad Sci U S A. 1984;81(12):3756–60.CrossRefPubMedPubMedCentral

24.

Mizuno H, Kitada K, Nakai K, Sarai A. PrognoScan: a new database for meta-analysis of the prognostic value of genes. BMC Med Genet. 2009;2:18.

25.

Pawitan Y, Bjohle J, Amler L, Borg AL, Egyhazi S, Hall P, Han X, Holmberg L, Huang F, Klaar S, et al. Gene expression profiling spares early breast cancer patients from adjuvant therapy: derived and validated in two population-based cohorts. Breast Cancer Res. 2005;7(6):R953–64.CrossRefPubMedPubMedCentral

26.

Ross SR. Mouse mammary tumor virus molecular biology and oncogenesis. Viruses. 2010;2(9):2000–12.CrossRefPubMedPubMedCentral

27.

Uhlen M, Zhang C, Lee S, Sjostedt E, Fagerberg L, Bidkhori G, Benfeitas R, Arif M, Liu Z, Edfors F, et al. A pathology atlas of the human cancer transcriptome. Science. 2017;357(6352):eaan2507. https://doi.org/10.1126/science.aan2507.

28.

Danielson KG, Knepper JE, Kittrell FS, Butel JS, Medina D, Durban EM. Clonal populations of the mouse mammary cell line, COMMA-D, which retain capability of morphogenesis in vivo. In Vitro Cell Dev Biol. 1989;25(6):535–43.CrossRefPubMed

29.

Hwang ES, Choi A, Ho IC. Transcriptional regulation of GATA-3 by an intronic regulatory region and fetal liver zinc finger protein 1. J Immunol. 2002;169(1):248–53.CrossRefPubMed

30.

Asselin-Labat ML, Sutherland KD, Barker H, Thomas R, Shackleton M, Forrest NC, Hartley L, Robb L, Grosveld FG, van der Wees J, et al. Gata-3 is an essential regulator of mammary-gland morphogenesis and luminal-cell differentiation. Nat Cell Biol. 2007;9(2):201–9.CrossRefPubMed

31.

Asselin-Labat ML. Mammary stem and progenitor cells: critical role of the transcription factor Gata-3. Med Sci (Paris). 2007;23(12):1077–80.CrossRef

32.

Dydensborg AB, Rose AA, Wilson BJ, Grote D, Paquet M, Giguere V, Siegel PM, Bouchard M. GATA3 inhibits breast cancer growth and pulmonary breast cancer metastasis. Oncogene. 2009;28(29):2634–42.CrossRefPubMed

33.

Hsu KS, Kao HY. PML: regulation and multifaceted function beyond tumor suppression. Cell & bioscience. 2018;8:5.CrossRef

34.

Dundr M. Nuclear bodies: multifunctional companions of the genome. Curr Opin Cell Biol. 2012;24(3):415–22.CrossRefPubMedPubMedCentral

35.

Scaglioni PP, Yung TM, Cai LF, Erdjument-Bromage H, Kaufman AJ, Singh B, Teruya-Feldstein J, Tempst P, Pandolfi PP. A CK2-dependent mechanism for degradation of the PML tumor suppressor. Cell. 2006;126(2):269–83.CrossRefPubMed

36.

Sarno S, Reddy H, Meggio F, Ruzzene M, Davies SP, Donella-Deana A, Shugar D, Pinna LA. Selectivity of 4,5,6,7-tetrabromobenzotriazole, an ATP site-directed inhibitor of protein kinase CK2 ('casein kinase-2′). FEBS Lett. 2001;496(1):44–8.CrossRefPubMed

37.

Hofmann TG, Will H. Body language: the function of PML nuclear bodies in apoptosis regulation. Cell Death Differ. 2003;10(12):1290–9.CrossRefPubMed

38.

Meek DW, Cox M. Induction and activation of the p53 pathway: a role for the protein kinase CK2? Mol Cell Biochem. 2011;356(1–2):133–8.CrossRefPubMed

39.

Scaglioni PP, Yung TM, Choi S, Baldini C, Konstantinidou G, Pandolfi PP. CK2 mediates phosphorylation and ubiquitin-mediated degradation of the PML tumor suppressor. Mol Cell Biochem. 2008;316(1–2):149–54.CrossRefPubMed

40.

Ehm P, Nalaskowski MM, Wundenberg T, Jucker M. The tumor suppressor SHIP1 colocalizes in nucleolar cavities with p53 and components of PML nuclear bodies. Nucleus. 2015;6(2):154–64.CrossRefPubMedPubMedCentral

41.

Hwang ES, Ho IC. Regulation of thymocyte homeostasis by Fliz1. Immunology. 2002;106(4):464–9.CrossRefPubMedPubMedCentral

42.

Kalathur M, Toso A, Chen J, Revandkar A, Danzer-Baltzer C, Guccini I, Alajati A, Sarti M, Pinton S, Brambilla L, et al. A chemogenomic screening identifies CK2 as a target for pro-senescence therapy in PTEN-deficient tumours. Nat Commun. 2015;6:7227.CrossRefPubMed

43.

Grande MA, van der Kraan I, van Steensel B, Schul W, de The H, van der Voort HT, de Jong L, van Driel R. PML-containing nuclear bodies: their spatial distribution in relation to other nuclear components. J Cell Biochem. 1996;63(3):280–91.CrossRefPubMed

44.

Sun J, Xu H, Subramony SH, Hebert MD. Interactions between coilin and PIASy partially link Cajal bodies to PML bodies. J Cell Sci. 2005;118(Pt 21):4995–5003.CrossRefPubMed

45.

Fu M, Blackshear PJ. RNA-binding proteins in immune regulation: a focus on CCCH zinc finger proteins. Nat Rev Immunol. 2017;17(2):130–43.CrossRefPubMed

46.

Hurt JA, Obar RA, Zhai B, Farny NG, Gygi SP, Silver PA. A conserved CCCH-type zinc finger protein regulates mRNA nuclear adenylation and export. J Cell Biol. 2009;185(2):265–77.CrossRefPubMedPubMedCentral

47.

Anwar D, Takahashi H, Watanabe M, Suzuki M, Fukuda S, Hatakeyama S. p53 represses the transcription of snRNA genes by preventing the formation of little elongation complex. Biochim Biophys Acta. 2016;1859(8):975–82.CrossRefPubMed

48.

Cerami E, Gao J, Dogrusoz U, Gross BE, Sumer SO, Aksoy BA, Jacobsen A, Byrne CJ, Heuer ML, Larsson E, et al. The cBio cancer genomics portal: an open platform for exploring multidimensional cancer genomics data. Cancer discovery. 2012;2(5):401–4.CrossRefPubMed

49.

Albergaria A, Paredes J, Sousa B, Milanezi F, Carneiro V, Bastos J, Costa S, Vieira D, Lopes N, Lam EW, et al. Expression of FOXA1 and GATA-3 in breast cancer: the prognostic significance in hormone receptor-negative tumours. Breast Cancer Res. 2009;11(3):R40.CrossRefPubMedPubMedCentral

50.

Ciocca V, Daskalakis C, Ciocca RM, Ruiz-Orrico A, Palazzo JP. The significance of GATA3 expression in breast cancer: a 10-year follow-up study. Hum Pathol. 2009;40(4):489–95.CrossRefPubMed

51.

Demir H, Turna H, Can G, Ilvan S. Clinicopathologic and prognostic evaluation of invasive breast carcinoma molecular subtypes and GATA3 expression. J BUON. 2010;15(4):774–82.PubMed

52.

Fang SH, Chen Y, Weigel RJ. GATA-3 as a marker of hormone response in breast cancer. J Surg Res. 2009;157(2):290–5.CrossRefPubMed

53.

Voduc D, Cheang M, Nielsen T. GATA-3 expression in breast cancer has a strong association with estrogen receptor but lacks independent prognostic value. Cancer Epidemiol Biomark Prev. 2008;17(2):365–73.CrossRef

Title: Regulation of the oncogenic phenotype by the nuclear body protein ZC3H8
Authors: John A. Schmidt
Keith G. Danielson
Emily R. Duffner
Sara G. Radecki
Gerard T. Walker
Amber Shelton
Tianjiao Wang
Janice E. Knepper
Publication date: 01-12-2018
Publisher: BioMed Central
Published in: BMC Cancer / Issue 1/2018
Electronic ISSN: 1471-2407
DOI: https://doi.org/10.1186/s12885-018-4674-1

Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras

Prof. Fabrice Barlesi

Developed by: Springer Medicine

Keynote webinar | Spotlight on medication adherence

Springer Medicine

Abstract

Background

Methods

Results

Conclusions

Please log in to get access to this content

Other articles of this Issue 1/2018

Chemotherapy weakly contributes to predicted neoantigen expression in ovarian cancer

A prospective clinical and biological database for pancreatic adenocarcinoma: the BACAP cohort

A rare case of primary breast angiosarcoma in a male: a case report

Pathological and molecular diagnosis of bilateral inguinal lymph nodes metastases from low-grade endometrial adenocarcinoma: a case report with review of the literature

Sidedness and TP53 mutations impact OS in anti-EGFR but not anti-VEGF treated mCRC - an analysis of the KRAS registry of the AGMT (Arbeitsgemeinschaft Medikamentöse Tumortherapie)

Loss of PDPK1 abrogates resistance to gemcitabine in label-retaining pancreatic cancer cells

Keynote webinar | Spotlight on antibody–drug conjugates in cancer