Differential expression of endogenous plant cell wall degrading enzyme genes in the stick insect (Phasmatodea) midgut

Abstract

BackgroundStick and leaf insects (Phasmatodea) are an exclusively leaf-feeding order of insects with no record of omnivory, unlike other “herbivorous” Polyneoptera. They represent an ideal system for investigating the adaptations necessary for obligate folivory, including plant cell wall degrading enzymes (PCWDEs). However, their physiology and internal anatomy is poorly understood, with limited genomic resources available.ResultsWe de novo assembled transcriptomes for the anterior and posterior midguts of six diverse Phasmatodea species, with RNA-Seq on one exemplar species, Peruphasma schultei. The latter’s assembly yielded >100,000 transcripts, with over 4000 transcripts uniquely or more highly expressed in specific midgut sections. Two to three dozen PCWDE encoding gene families, including cellulases and pectinases, were differentially expressed in the anterior midgut. These genes were also found in genomic DNA from phasmid brain tissue, suggesting endogenous production. Sequence alignments revealed catalytic sites on most PCWDE transcripts. While most phasmid PCWDE genes showed homology with those of other insects, the pectinases were homologous to bacterial genes.ConclusionsWe identified a large and diverse PCWDE repertoire endogenous to the phasmids. If these expressed genes are translated into active enzymes, then phasmids can theoretically break plant cell walls into their monomer components independently of microbial symbionts. The differential gene expression between the two midgut sections provides the first molecular hints as to their function in living phasmids. Our work expands the resources available for industrial applications of animal-derived PCWDEs, and facilitates evolutionary analysis of lower Polyneopteran digestive enzymes, including the pectinases whose origin in Phasmatodea may have been a horizontal transfer event from bacteria.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-917) contains supplementary material, which is available to authorized users.

Cloning, expression and characterization of a cold-adapted endo-1, 4-β-glucanase from Citrobacter farmeri A1, a symbiotic bacterium of Reticulitermes labralis

“…Although many endoglucanases from various microorganisms including fungus, bacteria, and actinomycetes have been studied, no endoglucanase from C. farmeri has been characterized to date (Shelomi et al, 2014; Asgher, Ahmad & Iqbal, 2013). In the present study, a cold-adapted endoglucanase was cloned from C. farmeri A1 and expressed in E. coli .…”

“…Cellulases, secreted by bacteria, fungi, and invertebrates, degrade β -1,4 glycosidic bonds of cellulose into sugars (Ogura et al, 2006; Limayem & Ricke, 2012; Shelomi et al, 2014). The complete degradation of cellulose requires three different cellulases: endo-1,4- β -glucanase, exo-1,4- β -glucanase, and 1,4- β –glucosidase (Ozioko, Eze & Chilaka, 2013).…”

De novo transcriptome analysis of the excretory tubules of Carausius morosus (Phasmatodea) and possible functions of the midgut ‘appendices’

“…The abundance of calcium-regulatory transcripts there matches what is known about Phasmatodea MpgTs and their relationship to calcium [50, 51]. The transcriptome also identified the known digestive enzymes in the midgut, including the Phasmatodea-specific pectinases [49], suggesting sufficient coverage. That the majority of identified transcripts had closest sequence homology to other Polyneoptera insect genes suggests low contamination and little chance that these are not endogenous Carausius genes.…”

“…I searched for biomineralization genes based on the listing of Livingston et al [48], due to the hypothesized existence in the AoMs of carbonic anhydrase [33] and otopetrin [49], and the known role of the Phasmatodea MpgTs in calcium elimination and deposition of unique calcium phosphate and/or calcium oxalate layers on the eggs [50–52]. These include bone morphogenic proteins, collagen, and regucalcin (one of which was highly and differentially expressed in the MpgTs, another in the AoMs), and matrix proteases (Fig 6).…”

“…The effects of this alkalinization could be to counteract or disable midgut digestive enzymes, but it may also have functions within the AoM tubules and ampules themselves. Once again identified in the AoMs [49] is high and differential expression of the otolith formation and biomineralization gene, otopetrin [62] (Fig 6). The two otopetrins differentially expressed in the AoMs (Carausius_C5193 & Carausius_C8672) have the 1 st and 7th greatest log2 fold change recorded between the AoM and MpgT (S1 Table).…”