Molecular cloning and characterization of a short peptidoglycan recognition protein from silkworm Bombyx mori

Area of Science:

• Insect immunity
• Molecular biology
• Biochemistry

Background:

• Peptidoglycans are key bacterial components triggering insect innate immunity.
• Peptidoglycan Recognition Proteins (PGRPs) are pattern-recognition receptors modulating immune responses.
• Some PGRPs possess enzymatic activity to degrade peptidoglycans, while others function solely as receptors.

Purpose of the Study:

• To clone and functionally analyze PGRP-S4, a short-form PGRP from the silkworm Bombyx mori.
• To investigate the immune functions and molecular mechanisms of PGRP-S4.

Main Methods:

• Gene cloning and protein expression of Bombyx mori PGRP-S4.
• Analysis of PGRP-S4 expression patterns in response to bacterial challenges.
• Biochemical assays to assess binding to bacteria and peptidoglycans, enzymatic activity, and effects on bacterial growth.
• Scanning electron microscopy to visualize bacterial cell surface disruption.
• Measurement of prophenoloxidase activation.

Main Results:

• PGRP-S4 gene encodes a 199-amino acid protein expressed in fat body, haemocytes, and midgut.
• Expression of PGRP-S4 was significantly induced by bacterial challenges in the midgut.
• Recombinant PGRP-S4 demonstrated binding to bacteria and peptidoglycans, inhibited bacterial growth, and hydrolyzed E. coli peptidoglycan in the presence of Zn2+.
• PGRP-S4 disrupted bacterial cell surfaces and enhanced peptidoglycan-induced prophenoloxidase activation.

Conclusions:

• Bombyx mori PGRP-S4 exhibits multiple immune functions, including direct antibacterial activity and modulation of the innate immune response.
• PGRP-S4 acts as both a pattern-recognition receptor and an enzyme, contributing to silkworm defense against bacterial infections.