454-hypoxia-bruchid
Transcriptome analysis of the cowpea bruchid: Oxygen deprivation alters gene expression of metabolic enzymes and heat shock proteins
Oxygen is of fundamental importance for many living organisms including insects.  Hermetic storage uses airtight storage facilities to withhold the oxygen required for insect development, thus preventing damage by insect pests in stored grain.  It has been shown previously that cowpea bruchids (Callosobruchus maculatus) cease feeding and growth when the oxygen level drops to 3-6%.  However, although population expansion is temporarily arrested, a portion of larvae can survive extended periods of time hypoxia once brought back to normoxic condition.  To begin to understand fundamental mechanisms that enable insects to cope with oxygen deprivation, an ability rarely found in mammals, we constructed a 3'-anchored cDNA library and performed 454-pyrosequencing.  Quality filtering and contig assembly resulted in 20,846 unique sequences. Of these, 5,335 sequences (25.6%) had hits in BLASTX search (E =10-6), constituting a 2,979 unigene set.  Further analysis based on the Gene Ontology terms indicated that 1,036 genes were involved in diverse cellular functions.  Putative genes encoding glycolytic and TCA cycle enzymes as well as components of respiratory chain complexes were selected and assessed for transcript responses to low oxygen.  Majority of these genes are down-regulated, suggesting hypoxia represses metabolic activity.  Promoter analyses of representative heat shock protein (HSP) genes suggest the involvement of hypoxia-inducible transcription factor 1 (HIF1) involvement in regulating these hypoxia-induced genes.  This activator function has been confirmed by transient co-transfection in S2 cells of HIF1 subunits and the HSP promoter-driven reporter.