Insect group II chitinase OfChtII promotes chitin degradation during larva–pupa molting

Abstract

The insect group II chitinase (ChtII, also known as Cht10) is a unique chitinase with multiple catalytic and chitin-binding domains. It has been proven genetically to be an essential chitinase for molting. However, ChtII's role in chitin degradation during insect development remains poorly understood. Obtaining this knowledge is the key to fully understanding the chitin degradation system in insects. Here, we investigated the role of OfChtII during the molting of Ostrinia furnacalis, a model lepidopteran pest insect. OfChtII was expressed earlier than OfChtI (OfCht5) and OfChi-h, at both the gene and protein levels during larva-pupa molting as evidenced by quantitative polymerase chain reaction and western blot analyses. A truncated OfChtII, OfChtII-B4C1, was recombinantly expressed in Pichia pastoris cells and purified to homogeneity. The recombinant OfChtII-B4C1 loosened compacted chitin particles and produced holes in the cuticle surface as evidenced by scanning electron microscopy. It synergized with OfChtI and OfChi-h when hydrolyzing insoluble α-chitin. These findings suggested an important role for ChtII during insect molting and also provided a strategy for the coordinated degradation of cuticular chitin during insect molting by ChtII, ChtI and Chi-h.

Systematic identification and characterization of long noncoding RNAs (lncRNAs) during Aedes albopictus development

“…Functional annotation of the pink, magenta and salmon modules indicated that enriched GO terms and KEGG pathways were significantly associated with metabolism, coinciding with rapid metamorphic development during these stages. For instance, the enrichment of chitin metabolic processes and chitin binding are closely related to the formation and degradation of chitin, which is a major structural component of the insect cuticle that protects insects from chemical erosion, physical abrasion and pathogenic invasion [ 53 , 54 ] and serves as an attachment matrix for other cuticular proteins, plays an important role in molting during the larval-larval and larval-pupal stages [ 55 , 56 ].…”

The DOMON domain protein LmKnk contributes to correct chitin content, pore canal formation and lipid deposition in the cuticle of Locusta migratoria during moulting

“…To accommodate body growth during development, insect cuticle must undergo periodic moulting by replacing the old cuticle by a new one (Moussian, 2010). Moulting can be subdivided in three phases: The preparatory phase including apolysis (i.e., detachment of the old cuticle from the body surface), the cuticle induction phase when a new cuticle is formed and the ecdysial phase when the insect emerges from the old cuticle and the new cuticle maturates (Truman, 2019;Qu et al, 2021).…”

High-Speed Atomic Force Microscopy Reveals Factors Affecting the Processivity of Chitinases during Interfacial Enzymatic Hydrolysis of Crystalline Chitin

“…OfChtII possesses multiple CBM14 domains and GH18 domains. Here, a truncated form of OfChtII containing three CBM14 domains at the N-terminal and a GH18 domain at the C-terminal was used. , OfChi-h possesses an N-terminal fibronectin III domain and a C-terminal GH18 domain . All recombinant chitinases were secreted into the culture supernatant and were purified by a combination of ammonium sulfate precipitation and immobilized metal ion affinity chromatography (IMAC), as described previously. ,, All purified enzymes were dissolved in the sample buffer (50 mM sodium phosphate, 150 mM NaCl, pH 6.0) before use.…”

“…In chitin-containing organisms and some microbes that utilize chitin as a nutrient resource, three or more glycoside hydrolase family 18 (GH18) chitinases constitute an enzyme cocktail for chitin hydrolysis. , Synergism among the cocktail enzymes has been reported to improve hydrolytic efficiency in several bacteria, , fungi , and insects. , Current research studies suggest that the synergism attributes to the different and complementary hydrolytic pattern of GH18 chitinases . In the most well-studied systems from the bacterium Serratia marcescens, the synergy is known to take place between three GH18 chitinases, SmChiA, SmChiB, and SmChiC. ,, Both SmChiA and SmChiB are processive enzymes. , The directionality of the processive motion is linked to the architecture of the substrate-binding sites of a processive enzyme, which, through the distribution of binding energies per site, provide directionality to the (partial) dissociation and rebinding step that needs to happen in between each processive cut .…”