title: Different functions of the insect soluble and membrane-bound trehalase genes in chitin biosynthesis revealed by RNA interference.
abstract: BACKGROUND
  Trehalase, an enzyme that hydrolyzes trehalose to yield two glucose molecules, plays a pivotal role in various physiological processes. In recent years, trehalase proteins have been purified from several insect species and are divided into soluble (Tre-1) and membrane-bound (Tre-2) trehalases. However, no functions of the two trehalases in chitin biosynthesis in insects have yet been reported.
  
  
  RESULTS
  The membrane-bound trehalase of Spodoptera exigua (SeTre-2) was characterized in our laboratory previously. In this study, we cloned the soluble trehalase gene (SeTre-1) and investigated the tissue distribution and developmental expression pattern of the two trehalase genes. SeTre-1 was expressed highly in cuticle and Malpighian tubules, while SeTre-2 was expressed in tracheae and fat body. In the midgut, the two trehalase genes were expressed in different locations. Additionally, the expression profiles of both trehalase mRNAs and their enzyme activities suggest that they may play different roles in chitin biosynthesis. The RNA interference (RNAi) of either SeTre-1 or SeTre-2 was gene-specific and effective, with efficiency rates up to 83% at 72 h post injection. After RNAi of SeTre-1 and SeTre-2, significant higher mortality rates were observed during the larva-pupa stage and pupa-adult stage, and the lethal phenotypes were classified and analyzed. Additionally, the change trends of concentration of trehalose and glucose appeared reciprocally in RNAi-mutants. Moreover, knockdown of SeTre-1 gene largely inhibited the expression of chitin synthase gene A (CHSA) and reduced the chitin content in the cuticle to two-thirds relative to the control insects. The chitin synthase gene B (CHSB) expression, however, was inhibited more by the injection of dsRNA for SeTre-2, and the chitin content in the midgut decreased by about 25%.
  
  
  CONCLUSIONS
  SeTre-1 plays a major role in CHSA expression and chitin synthesis in the cuticle, and SeTre-2 has an important role in CHSB expression and chitin synthesis in the midgut.
authors.affiliation: State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Chen
authors.forename: Bin
authors.initials: B
authors.lastname: Tang
authors.forename: Hongxin
authors.initials: H
authors.lastname: Chen
authors.forename: Qiong
authors.initials: Q
authors.lastname: Yao
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Huang
authors.forename: Jing
authors.initials: J
authors.lastname: Chen
authors.forename: Daowei
authors.initials: D
authors.lastname: Zhang
authors.forename: Wenqing
authors.initials: W
authors.lastname: Zhang
mesh_terms: D000818:Animals; D002686:Chitin / Q000096:biosynthesis*; D002687:Chitin Synthase / Q000235:genetics; D020869:Gene Expression Profiling; D007313:Insecta; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D012995:Solubility; D018411:Spodoptera / Q000201:enzymology / Q000235:genetics; D014018:Tissue Distribution; D014198:Trehalase / Q000235:genetics*
chemical_list: D034741:RNA, Small Interfering; D002686:Chitin; D002687:Chitin Synthase; D014198:Trehalase
references.citation: BMC Mol Biol. 2008 May 20;9:51
references.pmid: 18492231
references.citation: Insect Biochem Mol Biol. 2005 Jun;35(6):515-27
references.pmid: 15857758
references.citation: Comp Biochem Physiol B Biochem Mol Biol. 2008 Mar;149(3):447-53
references.pmid: 18178495
references.citation: Peptides. 2006 Jun;27(6):1226-34
references.pmid: 16386334
references.citation: Protein Expr Purif. 2009 Jun;65(2):185-9
references.pmid: 19073263
references.citation: Biochem J. 1992 Nov 15;288 ( Pt 1):19-22
references.pmid: 1445264
references.citation: Insect Biochem Mol Biol. 2007 May;37(5):409-17
references.pmid: 17456436
references.citation: Insect Biochem Mol Biol. 2008 Aug;38(8):788-95
references.pmid: 18625402
references.citation: Science. 1961 Jul 7;134(3471):54-5
references.pmid: 13694011
references.citation: Insect Biochem Mol Biol. 2008 Apr;38(4):452-66
references.pmid: 18342250
references.citation: Insect Mol Biol. 2005 Oct;14(5):453-63
references.pmid: 16164601
references.citation: J Insect Physiol. 2008 Feb;54(2):351-7
references.pmid: 18023454
references.citation: Mol Microbiol. 1996 Jun;20(6):1287-96
references.pmid: 8809779
references.citation: J Insect Physiol. 2009 Nov;55(11):997-1002
references.pmid: 19615372
references.citation: J Exp Biol. 2003 Dec;206(Pt 24):4393-412
references.pmid: 14610026
references.citation: Proc Natl Acad Sci U S A. 2008 May 6;105(18):6650-5
references.pmid: 18436642
references.citation: J Comp Physiol B. 1996;166(1):1-10
references.pmid: 8621836
references.citation: Experientia. 1996 May 15;52(5):433-9
references.pmid: 8706810
references.citation: Biochem Biophys Res Commun. 2009 May 22;383(1):98-102
references.pmid: 19336219
references.citation: Insect Mol Biol. 2009 Nov;18(6):769-84
references.pmid: 19843188
references.citation: Insect Biochem Mol Biol. 2006 Jan;36(1):1-9
references.pmid: 16360944
references.citation: Tissue Cell. 1985;17(4):539-51
references.pmid: 18620141
references.citation: Zoolog Sci. 2008 Jul;25(7):693-8
references.pmid: 18828655
references.citation: Insect Mol Biol. 2005 Oct;14(5):501-8
references.pmid: 16164606
references.citation: Mech Dev. 2007 Mar;124(3):180-9
references.pmid: 17258436
references.citation: Biochim Biophys Acta. 1993 May 28;1173(2):217-24
references.pmid: 7916633