Longhorned beetles (Cerambycidae) are the most diverse group of predominantly wood‐feeding (xylophagous) insects on Earth. Larvae of most species feed within tissues of plants made up of large amounts of plant cell wall (PCW), which is notoriously difficult to digest. To efficiently access nutrients from their food source, cerambycid larvae have to deconstruct PCW polysaccharides – such as cellulose, hemicelluloses and pectin – requiring them to possess a diversity of plant cell wall degrading enzymes (PCWDEs) in their digestive tract. Genomic data for Cerambycidae are mostly limited to notorious forest pests and are lacking for most of the taxonomic groups. Consequently, our understanding of the distribution and evolution of cerambycid PCWDEs is quite limited. We addressed the numbers, kinds and evolution of cerambycid PCWDEs by surveying larval midgut transcriptomes from 23 species representing six of the eight recognized subfamilies of Cerambycidae and each with very diverse host types (i.e., gymnosperms, angiosperms, xylem, phloem, fresh or dead plant tissues). Using these data, we identified 340 new putative PCWDEs belonging to ten carbohydrate active enzyme families, including two gene families (GH7 and GH53) not previously reported from insects. The remarkably wide range of PCWDEs expressed by Cerambycidae should allow them to break down most PCW polysaccharides. Moreover, the observed distribution of PCWDEs encoded in cerambycid genomes agreed more with phylogenetic relationship of the species studied than with the taxonomic origin or quality of the host plant tissues.

The rise of functional diversity through gene duplication contributed to the adaption of organisms to various environments. Here we investigate the evolution of putative cellulases of the subfamily 2 of glycoside hydrolase family 5 (GH5_2) in the Cerambycidae (longhorned beetles), a megadiverse assemblage of mostly xylophagous beetles. Cerambycidae originally acquired GH5_2 from a bacterial donor through horizontal gene transfer (HGT), and extant species harbor multiple copies that arose from gene duplication. We ask how these digestive enzymes contributed to the ability of these beetles to feed on wood. We analyzed 113 GH5_2, including the functional characterization of 52 of them, derived from 25 species covering most subfamilies of Cerambycidae. Ancestral gene duplications led to five well-defined groups with distinct substrate specificity, allowing these beetles to break down, in addition to cellulose, polysaccharides that are abundant in plant cell walls (PCWs), namely, xyloglucan, xylan, and mannans. Resurrecting the ancestral enzyme originally acquired by HGT, we show it was a cellulase that was able to break down glucomannan and xylan. Finally, recent gene duplications further expanded the catalytic repertoire of cerambycid GH5_2, giving rise to enzymes that favor transglycosylation over hydrolysis. We suggest that HGT and gene duplication, which shaped the evolution of GH5_2, played a central role in the ability of cerambycid beetles to use a PCW-rich diet and may have contributed to their successful radiation.

With more than 36,000 species, the longhorned beetles (family Cerambycidae) are a mega‐diverse lineage of mostly xylophagous insects, all of which are represented by the sole sequenced genome of the Asian longhorned beetle (Anoplophora glabripennis; Lamiinae). Their successful radiation has been linked to their ability to degrade plant cell wall components using a range of so‐called plant cell wall‐degrading enzymes (PCWDEs). Our previous analysis of larval gut transcriptomes demonstrated that cerambycid beetles horizontally acquired genes encoding PCWDEs from various microbial donors; these genes evolved through multiple duplication events to form gene families. To gain further insights into the evolution of these gene families during the Cerambycidae radiation, we assembled draft genomes for four beetle species belonging to three subfamilies using long‐read nanopore sequencing. All the PCWDE‐encoding genes we annotated from the corresponding larval gut transcriptomes were present in these draft genomes. We confirmed that the newly discovered horizontally acquired glycoside hydrolase family 7 (GH7), subfamily 26 of GH43 (GH43_26), and GH53 (all of which are absent from the A. glabripennis genome) were indeed encoded by these beetles' genome. Most of the PCWDE‐encoding genes of bacterial origin gained introns after their transfer into the beetle genome. Altogether, we show that draft genome assemblies generated from nanopore long‐reads offer meaningful information to the study of the evolution of gene families in insects. We anticipate that our data will support studies aiming to better understand the biology of the Cerambycidae and other beetles in general.

Plants possess various defense strategies to counter attacks from microorganisms or herbivores. For example, plants reduce the cell-wall-macerating activity of pathogen- or insect-derived polygalacturonases (PGs) by expressing PG-inhibiting proteins (PGIPs). PGs and PGIPs belong to multi-gene families believed to have been shaped by an evolutionary arms race. The mustard leaf beetle Phaedon cochleariae expresses both active PGs and catalytically inactive PG pseudoenzymes. Previous studies demonstrated that (i) PGIPs target beetle PGs and (ii) the role of PG pseudoenzymes remains elusive, despite having been linked to the pectin degradation pathway. For further insight into the interaction between plant PGIPs and beetle PG family members, we combined affinity purification with proteomics and gene expression analyses, and identified novel inhibitors of beetle PGs from Chinese cabbage (Brassica rapa ssp. pekinensis). A beetle PG pseudoenzyme was not targeted by PGIPs, but instead interacted with PGIP-like proteins. Phylogenetic analysis revealed that PGIP-like proteins clustered apart from “classical” PGIPs but together with proteins, which have been involved in developmental processes. Our results indicate that PGIP-like proteins represent not only interesting novel PG inhibitor candidates in addition to “classical” PGIPs, but also fascinating new players in the arms race between herbivorous beetles and plant defenses.

Objectives : This study evaluated activities and ingredient contents concerning extracts according to extraction solvents of Insampaedok-san (IS, Renshen bai du-san). Methods : The herbal constituents of IS were extracted with water and 70% ethanol at 100°C for 2 hr. Using the HPLC system, the six ingredient contents of different solvent extracts of IS were analyzed. The nitric oxide (NO), prostaglandin E2 (PGE2) production and proinflammatory cytokines were measured in RAW264.7 cells stimulated with lipopolysaccharide (LPS). The macrophage-derived chemokine (MDC/CCL22) and regulated on activation normal T-cell expression and secreted (RANTES/CCL5) production were measured in HaCaT and BEAS-2B cells stimulated tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). The activities of glycerol-3-phosphate dehydrogenase (GPDH) and leptin level were measured in 3T3-L1 cells. Results : The calibration curves showed good linearity (r 2 =1.0000) for different concentration ranges. The contents of liquiritin, naringin, hesperidin, neohesperirin and glycyrrizin in 70% ethanol extracts of IS were relatively higher than that of water extract, however the content of ferulic acid in 70% ethanol and water extract of IS were similar. The extraction solvents of water and 70% ethanol were evaluated inhibitory effect on the production of NO, PGE2, TNF-α and IL-6 in RAW 264.7 cells. Their extractions were inhibitory effect on production of MDC/CCL22 and RANTES/CCL5 in HaCaT cell and BEAS-2B cell, respectively. In addition, evaluated reduced on GPDH activity and leptin level in 3T3-L1 preadipocyte cell. Conclusions : Our results suggest that IS extracts were inhibitory effects of disease such as inflammation, allergies and obesity.