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Abstract
Coxsackievirus B3 (CVB3) is a non-enveloped virus that has a single-stranded RNA genome. CVB3 induces myocarditis, and ultimately, dilated cardiomyopathy. A myocarditis variant of CVB3 (CVB3 H3) and its antibody-escape mutant (CVB3 10A1) were studied previously; H3 was found to induce myocarditis and 10A1 was found to be attenuated in infected mice. Although amino acid residue 165, located in a puff region of VP2, was found to be altered (i.e., the H3 asparagine was altered to aspartate in 10A1), the detailed mechanism of attenuation was not clearly elucidated. Here, DNA microarray technology was used to monitor changes in mRNA levels of infected mouse hearts after CVB3 H3 and 10A1 infection. This tool was used to elucidate the pathogenic mechanisms of viral infection by understanding virus-host interactions. We identified several genes, including protein tyrosine kinases, Ddr2 and Ptk2, as well as Clqb and Crry, involved in complement reactions, which may be involved in these viral processes. Thus, gene profiling can provide an opportunity to understand host immune responses to viral infection for gene therapy and may contribute to the identification of the target gene that is modified during treatment of viral myocarditis.
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Figure 1.
Characterization of CVB3 H3 and 10A1 infected mouse. (A) Histology of myocardial injury after CVB3 H3 and 10A1 infection. Injury to myocytes was identified with Evans Blue dye, and appears as a red image. Analysed hearts were harvested at 7 days after challenge. (B) Survival curve after CVB3 H3 and 10A1 challenge in mice. Each group contained 20 mice. The survival curve is shown as percentage survival against time post-infection.
Table 1.
Functional grouping of mRNAs of infected mouse heart that change more than 1.5 fold after CVB3 H3 and CVB3 10A1 infection
| Genea | FCb | |||
|---|---|---|---|---|
| 7 days pi | 13 days pi | |||
| H3 | 10A1 | H3 | 10A1 | |
| tyrosine kinase | ||||
| discoidin domain receptor family, member 2 | 2.0 | 2.1 | 2.0 | |
| PTK2 protein tyrosine kinase 2 | 2.0 | 2.1 | 2.5 | 2.7 |
| Eph receptor A5 | 1.6 | 1.6 | 1.8 | 2.3 |
| protein tyrosine phosphatase, receptor type, T | 2.2 | 1.8 | ||
| Kinase | ||||
| adenylate kinase 3 alpha-like | 2.1 | |||
| A kinase (PRKA) anchor protein 8 | 1.7 | 1.9 | 1.6 | 1.7 |
| calcium/calmodulin-dependent protein kinase kinase 1, alpha | 2.0 | 2.4 | ||
| mitogen activated protein kinase kinase kinase 1 | 1.8 | 2.3 | 1.7 | |
| mitogen activated protein kinase kinase kinase 4 | 2.3 | 2.4 | ||
| mitogen-activated protein kinase 11 | 1.5 | 1.6 | ||
| mitogen activated protein kinase 9 | 1.5 | 1.5 | 1.6 | |
| 3-phosphoinositide dependent protein kinase-1 | 1.9 | 2.1 | ||
| intracellular signaling cascade | ||||
| amyloid beta (A4) precursor protein-binding, family B, member 1 interacting protein | 2.6 | 2.0 | ||
| Rho guanine nucleotide exchange factor (GEF7) | 1.5 | 1.8 | 1.6 | |
| expressed sequence AW049765 | 1.6 | 2.0 | ||
| G protein-coupled receptor kinase 5 | 1.5 | 1.7 | ||
| growth factor receptor bound protein 14 | 1.6 | 1.8 | 2.0 | 2.4 |
| membrane interacting protein of RGS16 | 1.6 | 2.0 | ||
| neuromedin | 1.7 | 2.1 | ||
| phosphatidylinositol 3-kinase, catalytic, beta polypeptide | 2.0 | 2.1 | ||
| phosphatidylinositol 3-kinase, regulatory subunit, polypeptide 1 (p85 alpha) | 1.8 | 1.6 | ||
| phosphatidylinositol-4-phosphate 5-kinase, type 1 alpha | 1.7 | 2.5 | 2.5 | |
| protein phosphatase 1A, magnesium dependent, alpha isoform | 1.6 | 1.9 | 1.5 | |
| presenilin 1 | 2.1 | 1.8 | ||
| presenilin 2 | 2.1 | 1.8 | ||
| SH2 domain containing 3C | 2.2 | 2.6 | 2.5 | 2.7 |
| signal-induced proliferation associated gene 1 | 1.5 | 2.8 | 2.3 | |
| solute carrier family 21 (organic anion transporter), member 5 | 1.6 | 1.9 | 1.9 | |
| solute carrier family 25 (mitochondrial carrier; peroxisomal membrane protein), member 17 | 1.7 | 1.6 | ||
| solute carrier family 31, member 1 | 1.7 | 1.9 | 2.2 | 2.7 |
| solute carrier family 9 (sodium/hydrogen exchanger), member 1 | 1.8 | 2.0 | ||
| syntaxin 8 | 1.7 | 2.0 | ||
| thromboxane A synthase 1, platelet | 2.1 | 2.0 | ||
| tripartite motif protein 28 | 1.6 | 1.7 | 1.6 | 1.7 |
| immune response | ||||
| complement component 1, q subcomponent, beta polypeptide | 1.7 | 2.0 | 2.4 | 2.9 |
| chemokine (C-C motif) ligand 25 | 2.5 | 2.6 | ||
| CD8 antigen, alpha chain | 1.8 | 1.7 | ||
| complement component factor h | 2.1 | 2.0 | ||
| complement receptor related protein | 1.7 | 1.8 | ||
| colony stimulating factor 2 receptor, beta 1, low-affinity | 1.5 | 1.9 | ||
| interferon (alpha and beta) receptor 1 | 1.8 | 1.7 | ||
| interferon gamma receptor | 1.6 | 1.7 | ||
| lymphocyte antigen 6 complex, locus A | 1.7 | |||
| N-myc (and STAT) interactor | 1.9 | 2.1 | ||
| pre-B-cell colony-enhancing factor | 1.8 | 2.0 | 1.7 | 2.1 |
| apoptosis | ||||
| activating transcription factor 5 | 1.7 | |||
| cytochrome c, somatic | 1.6 | 1.5 | 2.0 | 1.6 |
| tumor necrosis factor receptor superfamily, member 17 | 1.6 | 1.6 | ||
| superoxide dismutase 2, mitochodrial | 1.5 | 1.8 | ||
| wild-type p53-induced gene 1 | 1.5 | 2.1 | 2.1 | |
| Protein transport | ||||
| ATP-binding cassette, sub-family F (GCN20), member 2 | 1.6 | 1.5 | 1.7 | |
| ATPase, Ca++ transporting, plasma membrane 2 | 1.6 | 1.6 | 1.5 | |
| ADP-ribosylation factor 6 | 2.0 | 2.1 | ||
| amyloid beta (A4) precursor protein-binding, family A, member 2 | 1.8 | 2.0 | ||
| component of oligomeric golgi complex 2 | 1.6 | 2.2 | 2.5 | |
| carboxypeptidase X 1 (M14 family) | 1.6 | |||
| cytochrome P450, 2j6 | 1.5 | 2.1 | 2.1 | |
| DNA segment, Chr 11, ERATO Doi 18, expressed | 1.8 | 1.8 | ||
| FXYD domain-containing ion transport regulator 1 | 2.1 | 2.3 | ||
| laminin, beta 3 | 2.0 | 2.2 | ||
| glutamate receptor, ionotropic, kainate 1 | 1.6 | 1.8 | 2.1 | 2.6 |
| myosin IC | 1.8 | 1.8 | 2.0 | 2.2 |
| RAB33A, member of RAS oncogene family | 1.8 | |||
| tripartite motif protein 28 | 1.6 | 1.7 | 1.7 | |
| ubiquitin-like 4 | 1.6 | 1.8 | 1.8 | |
| vacuolar protein sorting 29 (S. pombe) | 2.0 | 2.0 | ||
| vacuolar protein sorting 4b (yeast) | 1.6 | |||
| G-protein coupled receptor protein signaling pathway | ||||
| coagulation factor II (thrombin) receptor-like 3 | 1.6 | |||
| membrane interacting protein of RGS16 | 2.0 | |||
| prostaglandin I receptor (IP) | 1.8 | 2.0 | ||
| regulator of G-protein signaling 11 | 1.5 | 1.6 | ||
Table 2.
mRNAs of infected mouse heart that change more than 1.5 fold at 7 day post-infection
| Gene Symbola | |
|---|---|
| Induction in both CVB3 H3 and CVB3 10A1 infected mouse heart | Akap8, Alas1, Arhgef7, Atp2b2, Bdnf, C1qb, C230035B04, Cog2, Comt, Crry, Csf2rb1, Cstb, Cycs, D130006K24Rik, D17Wsu94e, Dcx, Ddr2, Elovl1, Epha5, Evi2b, Fxyd1, Gba2, Gprk5, Grb14, Grcc2F, Grik1, Gtf2ird1, Hnrpd, Jdap1-pending, LOC223754, Lpin1, Map3k1, Mpg, Mrpl23, Mrps14, Myo1c, Ntan1, Pbef-pending, Pclo, Ppm1a, Ptk2, Rfc2, Sh2d3c, Slc30a4, Slc31a1, Snap25bp, Sod2, Spg4, Surb7, Tfam, Tmpo, Tnfsf12, Trim28, Tspyl, Ub14, Uros, Wrn, Zfh4-pending |
| Induction in only CVB3 10A1 infected mouse heart | Abcf2, Acas2, Adam8, Arcn1, B630003E05Rik, Bat4, Cav2, Cnih, Cpd1-pending, Cyp2j6, D4Wsu125e, D5Buc26e, Deb1, E430018M07Rik, Eif2s2, Entpd1, Es2el, Esp, Gfi1, Gnpat, Inpp5e, Klf15, Litaf, Mapk9, MGC25248, MGC28597, Mllt3, Mnt, Mpv17, Mrps22, Ndr1, Nktr, Nr2f6, Nudt5, Panx1, Pip5k1a, Rab14, Rpo1–1, Scd1, Sdf2, Sipa1, Slc21a5, Sorbs1, Spp1, Sptlc2, Suv39h1, Tagln2, Tcfeb, Umpk, Uty, Wig1, Zap3-pending |
| Induction in only CVB3 H3 infected mouse heart | G1rzfp-per, Gyg1, Hn1, Lamr1, Mtap4, Osp94, Pik3r1 |
Table 3.
mRNAs of infected mouse heart that change more than 1.5 fold at 13 day post-infection
| Gene Symbola | |
|---|---|
| Induction in both CVB3 H3 and CVB3 10A1 infected mouse heart | Abcf2, Acas2, Aco1, Actn4, Adam8, Akap8, Alas1, Amfr, Apacd-pending, Apba2, Apbb1ip-pending, Apc7-pending, Arf6, Asah2, Asb8, Atf5, AW049765, B630002E05Rik, Bace, Bat4, BC026401, Brd4, C130052I12Rik, C1qb, Camkk1, Caskin2-pending, Ccl25, Cd8a, Cfh, Cited1, Clstn1, Clybl, Cnot8, Cog2, Comt, Cops3, Cpd1-pending, Crry, Crybb1, Ctsh, Cycs, Cyp2j6, D11Ertd18e, D130006K24Rik, D17Wsu94e, D730049H07Rik, Dcx, Ddr2, Deb1, Dnajb9, E430007K15Rik, E430018M07Rik, Elovl1, Entpd1, Epha5, Es2el, Evi2b, Expi, Fgfr2, Fibp, Fosb, Fxyd1, G0s2, G1rzfp-pending, Gas41-pending, Gcat, Gfi1, Gh, Gnpat, Gpi1, Grb14, Grcc2f, Grik1, Gtf2ird1, Gus, Gzma, H2afy, Hip1r, Hirip5, Hnrpd, Hs1bp3-pending, Ifnar1, Ifngr, Itpr5, Lamb3, Lamr1, Lcat, Lcp1, Litaf, LOC207352, LOC216543, LOC227619, LOC23373, LOC270153, LOC270153, Lpin1, Ly6a, Madh1, Map3k4, Mapk11, Mapk9, MGC25248, MGC28597, MGC40859, Minpp1, Mir16-pending, Mnt, Mpg, Mpv17, Mrpl12, Mrpl17, Mrps14, Mrps22, Mrps30, Ms4a4d, Mta1, Myo1c, Myo7a, N4wbp4-pending, Ncl, Ndr1, Nkx2–2, Nmi, Nmu, Nthl1, Osp94, Ovca2-pendingm Panx1, Papss1, Pbef-pending, Pcm1, Pdpk1, Pik5k1a, Plscr3, Pou2f2, Prodh, Psen1, Psen2, Pstpip1, Ptgir, Ptk2, Ptprt, Pxf, Rab14, Rab33a, Rangnrf-pending, Rfc2, Rgs11, Rnasep2-pending, Rpl8, Sc5d, Sh2d3c, Sipa1, Slc221a5, Slc25a17, Slc31a, Slc9a1, Smap-pending, Snap25bp, Sorbs1, Spg4, Spp1, Srprb, Ssbp3, Stx8, Surb7, Suv39h1, Tagln2, Tbx2, Tbxas1, Tmpo, Tnfrsf17, Trim28, Tslpr, Tspyl, Vamp4, Vps29, Wdr10, Wig1, Xpo1, Yars, Zap3-pending, Zfpm2 |
| Induction in only CVB3 10A1 infected mouse heart | Al181838, Al317223, Al317395, Al645590, Al848594, Arhgef7, Ass1, Atp2a2, AW060733, B430109J1, Bdnf, C130038E, C1qtnf5, Camk2b, Cnih, Cstb, D16Ertd36, D5Buc26e, D6Ertd253, Epb4.1l3, Fbxo16, Galns, Gba2, Gng10, Klf1a, Klf15, LOC22375, MGC27915, Mllt3, Mtap4, Ngef, Nktr, Notch1, Npr3, Nr2f6, Nudel-pending, Pclc, Pik3r1, Pirb, Ppp1cb, Ppp3ca, Prkacb, Prkcm, Rad51l1, Rbbp9m Rnase1, Rpo1-1, Rsu1, Scd1, Scotin-pending, Skz1-pending, Slc22a4, Slc25a11, Sox5, Sox6, Sptlc2, Stra6, Stx6, Surf4, Tfam, Tnfsf9, Trim16, U3-55k-pending, Ubl4, Uty, YEAST3–1, Zap3-pending |
| Induction in only CVB3 H3 infected mouse heart | Adh1, Al848594, Ak3l, Akt2, Atp2b2, C230035B04, Cpxm, D3Ucla1, F2rl3, Glud, Map3k1, Ncl, Nr2f6, Nudc, Rce1, Stx4a, Uros, Vps4b, Wwp2-pending, Zak-pending, Zfp52 |
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