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Povelones Laboratory

Research Interests

pove-lab-logo-2Our main research interest is innate immune recognition and elimination of pathogens. Our work focuses on the interaction between mosquitoes and the animal and human pathogens they transmit. As the most species-rich group of animals on the planet occupying a vast array of ecological niches, insects are a fantastic example of the potency of innate defenses.

Spotlight on our work

Rather than passive or willing carriers of pathogenic organism, mosquitoes are actually amazing pathogen killers. Taking mosquito interactions with malaria parasites as an example, the vast majority of the parasites ingested when a mosquito bites a malarious person are attacked and eliminated before they can mount an infection in the mosquito. It is the few parasites that survive (even one is sufficient), that are ultimately responsible for disease transmission. Similar interactions occur between mosquitoes and the other pathogens they transmit, like canine heartworm (Dirofilaria) and arboviruses (Zika, Dengue, Yellow Fever, West Nile, and Chikungunya). 

Arthropod vectors such as mosquitoes, sand flies and ticks are responsible for transmission of a large number of animal and human diseases worldwide. Studying these organisms may reveal general insights about innate immune defense mechanisms as well as provide novel avenues for controlling the terrible diseases they spread.

Some of the questions we are addressing:

  • What is the basis of pathogen recognition by the mosquito innate immune system and how do some pathogens manage to escape?
  • What is the biochemical mechanism leading from innate recognition to pathogen killing?
  • How is mosquito complement regulated?
  • How does steroid hormone signaling regulate mosquito immunity?

The mosquito has multiple lines of defense against invading pathogens. One potent barrier is found in its blood, called hemolymph (Figure 1).

Figure1-ii

 

Parasites migrate through the gut epithelium in order to escape the harsh digestive conditions of the gut lumen. Here they come into contact with the hemolymph. At this point parasites are recognized and killed by lysis or melanization. Those that survive transform into oocysts. Within the cyst wall parasites replicate. Approximately two weeks later, oocysts burst each releasing thousands of sporozoites into the hemolymph where they migrate to the salivary gland to injected into the next when the mosquito takes its next blood meal.

The hemolymph barrier is formed by a network of proteins referred to as the complement-like pathway due to its similarities to the vertebrate complement pathway. Two leucine-rich repeat (LRR) containing proteins, LRIM1 and APL1C, are essential are essential components of the mosquito complement pathway. These proteins circulate in the mosquito hemolymph in a disulfide-bonded dimer (Figure 2).

Figure2-ii

If either LRIM1 or APL1C is silenced by RNAi, the complex is undetectable in the hemolymph and parasite survival is massively increased. Before parasites are killed, the complement C3-like protein TEP1 is localized on their surface, marking them for destruction. The LRIM1/APL1C complex physically interacts with a proteolytically processed and highly reactive form of TEP1 called TEP1cut. The enzyme(s) responsible for the generation of steady-state levels of TEP1cut are not known. The interaction stabilizes TEP1cut in the hemolymph and is required for its localization to parasites during midgut invasion. When the LRIM1/APL1C complex is knocked-down, TEP1 fails to localize and the invading parasites are not killed. This immune pathway leading to parasite killing could be an important cause of natural refractoriness in non-vector mosquitoes. We recently discovered that the LRIM1/APL1C complex can also interact with 3 other members of the TEP family. Two of these were previously characterized to contribute to mosquito antibacterial defense reactions. We found that one of these, TEP3, also functions in mosquito immune reactions against parasites.

Additionally, we have found that LRIM1 and APL1C are defining members of a protein family, collectively named LRIMs (pronounced L-rims). Bioinformatic searches using specific features shared between LRIM1 and APL1C has uncovered approximately 20 family members falling into four distinct sub-families in the mosquito species Anopheles gambiae, Aedes aegypti and Culex quinquefasciatus (Figure 3).

 Figure3-ii

 

This family is not found in any other organism. Given the central role of LRR proteins in host defense in plants and animals, we are currently investigating the hypothesis that the repertoire of LRIMs may help the mosquito neutralize diverse pathogens, including the agents of human and animal diseases that they transmit (Figure 4).

Figure4-ii

We have recently discovered a novel branch of the immune system that is specifically regulated by the steroid hormone 20-hydroxyecdysone (20E). 20E is produced transiently in adult female mosquitoes in response to blood feeding and has a well-characterized role in egg production (Figure 1)

20E Signaling

We found that the 20E also regulates the immune protein LRIM9. LRIM9 protein is strongly induced in the mosquito hemolymph after blood feeding and protects the mosquito from infection by malaria parasites. Preliminary analysis revealed that LRIM9 is not part of the mosquito complement-like pathway and the mechanism by which is restricts parasite development is not known.

We are currently investigating

  • the mechanism of how LRIM9 leads to lower infection by Plasmodium
  • whether 20E regulates immune pathways in the mosquito hemolymph or other tissues
Mosquitoes are essential for the canine heartworm, Dirofilaria immitis,life cycle serving as both an intermediate hosts and vectors. Transmission requires that ingested D. immitismicrofilariae develop into infective third-stage larvae (L3) over an approximately 2-week period. As it develops in the mosquito, D. immitis interacts with different tissues and immunological barriers. We are addressing the role of immune signaling pathways in controlling D. immitis infection in the model vector Aedes aegypti.
 
povelab summer 2015

 

Lab Members
Letitia Thompson, Povelones Lab Letitia Thompson, Laboratory technician

Sarah Sneed, Graduate student, IGG program

Sutopa Dwivedi, Postdoctoral researcher

Barbara Biney, Undergraduate researcher, College of Arts and Sciences, Class of 2018

 
Previous Lab Members
anotoniabass

Antonia Bass, Graduate student, MVP program

Antonia investigated complement regulation inAnopheles gambiae(Rotation student, Fall 2015)

kristinderfus

Kristin Derfus, PennVet Class of 2019

Kristin is investigating the genetics of susceptibility ofAedes aegyptimosquitoes to infection by canine heartwormDirofilaria immitis(Summer 2015)

Andrew Hart, Povelones Lab

Andrew Hart, SUIP student

Andrew studied the complement pathway in maleAnopheles gambiae(Summer 2015)

Jenny Kwok, Povelones Lab

Jenny Kwok, PennVet Class of 2017

Jenny studied how activation of the mosquito's immune signaling affects infection by canine heartwormDirofilaria immitis(Summer 2015)

Valeria Reyes Ruiz, Povelones Lab

Valeria Reyes Ruiz, Graduate student, MVP program, JoinedSunny Shin's laboratory.

Valeria studied the specificity of mosquito complement activation (Rotation student, Spring 2015)