International Journal of Industrial Entomology and Biomaterials

Korean Society of Sericultural Science (한국잠사학회)
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Detection of the expression of a Bombyx mori Atypical Protein Kinase C in BmPLV-Infected Larval Midgut

  • Cao, Jian (Institute of Life Science, Jiangsu University) ;
  • He, Yuanqing (Institute of Life Science, Jiangsu University) ;
  • Li, Guohui (Institute of Life Science, Jiangsu University) ;
  • Chen, Keping (Institute of Life Science, Jiangsu University) ;
  • Kong, Jie (Institute of Life Science, Jiangsu University) ;
  • Wang, Fenghua (Institute of Life Science, Jiangsu University) ;
  • Shi, Jing (Institute of Life Science, Jiangsu University) ;
  • Yao, Qin (Institute of Life Science, Jiangsu University)
  • Received : 2011.03.24
  • Accepted : 2011.05.28
  • Published : 2011.06.30
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Abstract

Protein kinase C (PKC) is involved in many cellular signaling pathways, it participates in many physiological processes, such as cell cycle, growth, proliferation, differentiation and apoptosis. To investigate the effect of PKC on the silkworm midgut tissue infection of Bombyx mori parvo-like virus (BmPLV), a B. mori atypical protein kinase C (BmaPKC) gene was cloned from larval midgut tissue, expressed in E. coli and purified. Additionally, the BmPLV susceptible silkworm strain and resistant silkworm strain were used to test the effect of the B. mori infection on BmPLV. The result showed that BmaPKC encodes a predicted 586 amino acid protein, which contains a C-terminal kinase domain and an N-terminal regulatory domain. The maximum expression amount of the soluble (His)6-tagged fusion protein was detected after 0.8 mmol/L IPTG was added and cultured at $21^{\circ}C$. The (His) 6-tagged fusion protein revealed about 73 kDa molecular weight which confirmed by western blot and mass spectrography. Furthermore BmaPKC protein were detected at 0-72 h post-infection in BmPLVinfected larval midgut tissue, western blot showed that as time went on, the expression of BmaPKC increased gradually in susceptible strain, the expression quantity on 72 h is 5 times of 0 h. However, in resistant strain, the expression quantity is slightly lower than susceptible strain. But no significant change in resistant strain was observed as time went on. The available data suggest that BmaPKC may involve in the regulation of BmPLV proliferation.

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References

  1. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72, 248-254. https://doi.org/10.1016/0003-2697(76)90527-3
  2. Dedos SG, Kaltofen S, Birkenbeil H (2008) Protein kinase A and C are ''Gatekeepers'' of capacitative $Ca^{2+}$ entry in the prothoracic gland cells of the silkworm, Bombyx mori. J Insect Physiol 54, 878-882. https://doi.org/10.1016/j.jinsphys.2008.03.010
  3. Dettwiler S, Rommelaere J, Nuesch JP (1999) DNA unwinding functions of minute virus of mice NS1 protein are modulated specifically by the lambda isoform of protein kinase C. J Virol 73, 7410-7420.
  4. Kim S, Gailite I, Moussian B, Luschnig S, Goette M, Fricke K, Honemann-Capito M, Grubmuller H, Wodarz A (2009) Kinase-activity-independent functions of atypical protein kinase C in Drosophila. J Cell Sci 122, 3759-3771. https://doi.org/10.1242/jcs.052514
  5. Kovac J, Oster H, Leitges M (2007) Expression of the atypical protein kinase C (aPKC) isoforms [iota]/[lambda] and [zeta] during mouse embryogenesis. Gene Exp Patt 7, 187-196. https://doi.org/10.1016/j.modgep.2006.07.002
  6. Lachmann S, Rommeleare J, Nuesch JPF (2003) Novel PKC {eta} is required to activate replicative functions of the major nonstructural protein NS1 of minute virus of mice. J Virol 77, 8048-8060. https://doi.org/10.1128/JVI.77.14.8048-8060.2003
  7. Li XH, Wu XF, Yue WF, Liu JM, Li GL, Miao YG (2006) Proteomic analysis of the silkworm (Bombyx mori L.) hemolymph during developmental stage. J Proteome Res 5, 2809-2814. https://doi.org/10.1021/pr0603093
  8. Melowic HR, Stahelin RV, Blatner NR, Tian W, Hayashi K, Altman A, Cho W (2007) Mechanism of diacylglycerolinduced membrane targeting and activation of protein kinase C e. J Biol Chem 282, 21467-21476 https://doi.org/10.1074/jbc.M700119200
  9. Moscat J, Diaz-Meco MT, Albert A, Campuzano S (2006) Cell signaling and function organized by PB1 domain interactions. Mol Cell 23, 631-640. https://doi.org/10.1016/j.molcel.2006.08.002
  10. Stahelin RV, Digman MA, Medkova M, Ananthanarayanan B, Melowic HR, Rafter JD, Cho W (2005) Diacylglycerolinduced membrane targeting and activation of protein kinase C. J Biol Chem 280, 19784-19793. https://doi.org/10.1074/jbc.M411285200
  11. Stahelin RV, Digman MA, Medkova M, Ananthanarayanan B, Rafter JD, Melowic HR, Cho W (2004) Mechanism of diacylglycerol- induced membrane targeting and activation of protein kinase Cdelta. J Biol Chem 279, 29501-29512. https://doi.org/10.1074/jbc.M403191200
  12. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76, 4350-4354. https://doi.org/10.1073/pnas.76.9.4350
  13. Uno T, Nakao A, Fujiwara Y, Katsurauma C, Nakada T, Itoh O (2006) Molecular cloning and expression of protein kinase C from Bombyx mori. Arch Insect Biochem Physiol 61, 65-76. https://doi.org/10.1002/arch.20098
  14. Wang JM, Li Q, Du GS, Lu JX, Zou SQ (2009) Significance and expression of atypical protein kinase C-iota in human hepatocellular carcinoma. J Surg Res 154, 143-149. https://doi.org/10.1016/j.jss.2008.05.036