Related Experiment Video
Updated: Apr 30, 2026
Pharmacologic Induction of Epidermal Melanin and Protection Against Sunburn in a Humanized Mouse Model
Published on: September 7, 2013
Manduca sexta proprophenoloxidase activating proteinase-3 (PAP3) stimulates melanization by activating proPAP3,
Yang Wang1, Zhiqiang Lu1, Haobo Jiang1
1Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater, OK 74078, USA.
Abstract:
Melanization participates in various insect physiological processes including antimicrobial immune responses. Phenoloxidase (PO), a critical component of the enzyme system catalyzing melanin formation, is produced as an inactive precursor prophenoloxidase (proPO) and becomes active via specific proteolytic cleavage by proPO activating proteinase (PAP). In Manduca sexta, three PAPs can activate proPOs in the presence of two serine proteinase homologs (SPH1 and SPH2). While the hemolymph proteinases (HPs) that generate the active PAPs are known, it is unclear how the proSPHs (especially proSPH1) are activated. In this study, we isolated from plasma of bar-stage M. sexta larvae an Ile-Glu-Ala-Arg-p-nitroanilide hydrolyzing enzyme that cleaved the proSPHs. This proteinase, PAP3, generated active SPH1 and SPH2, which function as cofactors for PAP3 in proPO activation. Cleavage of the purified recombinant proSPHs by PAP3 yielded 38 kDa bands similar in mobility to the SPHs formed in vivo. Surprisingly, PAP3 also can activate proPAP3 to stimulate melanization in a direct positive feedback loop. The enhanced proPO activation concurred with the cleavage activation of proHP6, proHP8, proPAP1, proPAP3, proSPH1, proSPH2, proPOs, but not proHP14 or proHP21. These results indicate that PAP3, like PAP1, is a key factor of the self-reinforcing mechanism in the proPO activation system, which is linked to other immune responses in M. sexta.
Insights
This study identifies a novel proteinase, PAP3, in Manduca sexta that activates key enzymes in the melanization cascade, revealing a positive feedback loop crucial for insect immunity.
Area of Science:
- Insect physiology
- Biochemistry
- Immunology
Background:
- Melanization is vital for insect physiology, including immune responses.
- Phenoloxidase (PO) activation is central to melanization, involving prophenoloxidase (proPO) and proPO activating proteinases (PAPs).
- The activation mechanism of proSPHs, cofactors in proPO activation, remained unclear in Manduca sexta.
Purpose of the Study:
- To identify and characterize the enzyme responsible for activating proSPHs in Manduca sexta.
- To elucidate the role of this enzyme in the prophenoloxidase (proPO) activation system.
- To investigate potential positive feedback mechanisms in melanization.
Main Methods:
- Isolation and characterization of an Ile-Glu-Ala-Arg-p-nitroanilide hydrolyzing enzyme from Manduca sexta plasma.
- Assay of the enzyme's ability to cleave and activate proSPHs and other pro-forms.
- Analysis of cleavage products using SDS-PAGE.
Main Results:
- A novel proteinase, designated PAP3, was isolated and shown to cleave proSPH1 and proSPH2.
- PAP3 activates proSPHs, which then act as cofactors for PAP3 in proPO activation.
- PAP3 also activates proPAP3, establishing a positive feedback loop that enhances proPO activation and melanization.
Conclusions:
- PAP3 is a key enzyme in the Manduca sexta proPO activation system, distinct from PAP1.
- PAP3 mediates a self-reinforcing mechanism in melanization through positive feedback.
- This system is interconnected with other immune responses in Manduca sexta.
More Related Videos
Related Concept Videos
01:07MAPK Signaling Cascades
02:23Abnormal Proliferation
01:19Pigmentation
Melanin occurs in two primary forms: eumelanin that provides black and brown pigment and pheomelanin that provides red color. Dark-skinned individuals produce more melanin than those with pale...