Dataset Information

Characterization of a novel Manduca sexta beta-1, 3-glucan recognition protein (?GRP3) with multiple functions.

ABSTRACT: Recognition of pathogens by insect pattern recognition receptors is critical to mount effective immune responses. In this study, we reported a new member (?GRP3) of the ?-1, 3-glucan recognition protein (?GRP) family from the tobacco hornworm Manduca sexta. Unlike other members of the M. sexta ?GRP family proteins, which contain an N-terminal small glucan binding domain and a C-terminal large glucanase-like domain, ?GRP3 is 40-45 residues shorter at the N-terminus and lacks the small glucan binding domain. The glucanase-like domain of ?GRP3 is most similar to that of M. sexta microbe binding protein (MBP) with 78% identity. ?GRP3 transcript was mainly expressed in the fat body, and both its mRNA and protein levels were not induced by microorganisms in larvae. Recombinant ?GRP3 purified from Drosophila S2 cells could bind to several Gram-negative and Gram-positive bacteria and yeast, as well as to laminarin (?-1, 3-glucan), mannan, lipopolysaccharide (LPS), lipoteichoic acid (LTA), and meso-diaminopimelic acid (DAP)-type peptidoglycan (PG), but did not bind to Lysine-type PG. Binding of ?GRP3 to laminarin could be competed well by free laminarin, mannan, LPS and LTA, but almost not competed by free PGs. Recombinant ?GRP3 could agglutinate Bacillus cereus and Escherichia coli in a calcium-dependent manner and showed antibacterial (bacteriostatic) activity against B. cereus, novel functions that have not been reported for the ?GRP family proteins before. M. sexta ?GRP3 may serve as an immune surveillance receptor with multiple functions.

SUBMITTER: Rao XJ

PROVIDER: S-EPMC4143429 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Characterization of a novel Manduca sexta beta-1, 3-glucan recognition protein (βGRP3) with multiple functions.

Rao Xiang-Jun XJ Zhong Xue X Lin Xin-Yu XY Huang Xiao-Hong XH Yu Xiao-Qiang XQ

Insect biochemistry and molecular biology 20140619

Recognition of pathogens by insect pattern recognition receptors is critical to mount effective immune responses. In this study, we reported a new member (βGRP3) of the β-1, 3-glucan recognition protein (βGRP) family from the tobacco hornworm Manduca sexta. Unlike other members of the M. sexta βGRP family proteins, which contain an N-terminal small glucan binding domain and a C-terminal large glucanase-like domain, βGRP3 is 40-45 residues shorter at the N-terminus and lacks the small glucan bind ...[more]

PMID: 24952171

Similar Datasets

Project description:Although the importance of peptidoglycan recognition proteins (PGRPs) in detecting bacteria and promoting immunity is well recognized in Drosophila melanogaster and other insect species, such a role has not yet been experimentally established for PGRPs in the tobacco hornworm, Manduca sexta. In this study, we purified M. sexta PGRP1 from the baculovirus-insect cell expression system, tested its association with peptidoglycans and intact bacteria, and explored its possible link with the prophenoloxidase activation system in larval hemolymph. Sequence comparison suggested that PGRP1 is not an amidase and lacks residues for interacting with the carboxyl group of meso-diaminopimelic acid-peptidoglycans (DAP-PGs). M. sexta PGRP1 gene was constitutively expressed at a low level in fat body, and the mRNA concentration became much higher after an injection of Escherichia coli. Consistently, the protein concentration in larval plasma increased in a time-dependent manner after the immune challenge. Purified recombinant PGRP1 specifically bound to soluble DAP-PG of E. coli but not to soluble Lys-type PG of Staphylococcus aureus. In addition, this recognition protein completely bound to insoluble PGs from Micrococcus luteus, Bacillus megaterium and Bacillus subtilis, whereas its association with the bacterial cells was low even though their peptidoglycans are exposed on the cell surface. After PGRP1 had been added to plasma of naïve larvae in the absence of microbial elicitor, there was a concentration-dependent increase in prophenoloxidase activation. Phenoloxidase activity, as usual, increased after the plasma was incubated with peptidoglycans or bacterial cells. These increases became more prominent when insoluble M. luteus or B. megaterium PG or soluble E. coli PG and PGRP1 were both present. Statistic analysis suggested a synergistic effect caused by interaction between PGRP1 and these PGs. Taken together, these results indicated that PGRP1 is a member of the M. sexta prophenoloxidase activation system, which recognizes peptidoglycans from certain bacteria and initiates the host defense response. The unexplained difference between the purified PGs and intact bacteria clearly reflects our general lack of understanding of PGRP1-mediated recognition and how it leads to proPO activation.

Project description:Detection of pathogenic invaders is the essential first step of a successful defense response in multicellular organisms. In this study, we have identified a new member of the β-1,3-glucanase-related protein superfamily from the tobacco hornworm Manduca sexta. This protein, designated microbe binding protein (MBP), is 61% identical in sequence to Bombyx mori Gram-negative bacteria binding protein, but only 34-36% identical to M. sexta β-1,3-glucan recognition protein-1 and 2. Its mRNA levels were strongly up-regulated in hemocytes and fat body of immune challenged larvae, along with an increase in concentration of the plasma protein. We expressed M. sexta MBP in a baculovirus-insect cell system. The purified protein associated with intact bacteria and fungi. It specifically bound to lipoteichoic acid, lipopolysaccharide, diaminopimelic acid-type peptidoglycans (DAP-PGs) from Escherichia coli and Bacillus subtilis, but less so to laminarin or Lys-type PG from Staphylococcus aureus. The complex binding pattern was influenced by other plasma factors and additional microbial surface molecules. After different amounts of MBP had been incubated with larval plasma on ice, a concentration-dependent increase in phenoloxidase (PO) activity occurred in the absence of any microbial elicitor. The activity increase was also observed in the mixture of plasma and a bacterial or fungal cell wall component. The prophenoloxidase (proPO) activation became more prominent when DAP-PGs, Micrococcus luteus Lys-PG, or lipoteichoic acid was included in the mixture of MBP and plasma. Statistic analysis suggested that a synergistic enhancement of proPO activation was caused by an interaction between MBP and these elicitors, but not S. aureus Lys-PG, lipopolysaccharide, curdlan, or laminarin. These data indicate that M. sexta MBP is a component of the surveillance mechanism and, by working together with other pattern recognition molecules and serine proteinases, triggers the proPO activation system.

Dataset Information

Characterization of a novel Manduca sexta beta-1, 3-glucan recognition protein (?GRP3) with multiple functions.

Publications

Characterization of a novel Manduca sexta beta-1, 3-glucan recognition protein (βGRP3) with multiple functions.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets