Day One

I am doing my research in the lab of Dr. Riehle. He is studying ways to prevent the transmission of Malaria by modifying genes that increase the activity of the Insulin Signaling Pathway(ISP). I'll explain that later; this is what I did today.

Right off the bat Dr. Riehle had me disecting mosquitos under a microscope. A mosquito has (I think) 6 or so sections in it's abdomen. The parts we are intersted in are the ovaries, the fat body, and the midgut. This said, we are only interested in disecting females(although I did a couple males just for giggles). The ovaries are obtained by ripping(with very cool, very expensive tweezers) the second section of the abdomen(from the back) from the rest of the body. The resulting mess is eight white tubules(they kind of look like the arms of a sea anemone) and the ovaries. The white tubules are part of the fat body(the fat body is essentially anything that isn't the epithilial tissue or ovaries that is in the abdomen). The midgut is in the remaining sections of the abdomen, and is surrounded by fat body. After practicing my disection abilities, I was introduced to the grad student who will be mentoring me, Anam Aric.

Anam took me out of the microscope room and back into the main lab. She started telling me the basics of the mosquito life cycle, reproduction, etc. and the basic ideas behind using the Insulin Signaling Pathway activity to control the lifespan and reproductive capabilities of the mosquito. First off, I will describe the two different diseases and the mosquito each is associated with.

The first is Anapholes Stephensi, which is a less evolved mosquito that is known to transmit malaria. Malaria is a parasite(not a bacteria or a virus) that develops inside the epithilial tissue(the tissue that lines the midgut). The male and female gamates form inside this tissue and eventually burst out and make their way to the salvatory glands. It can take up to two weeks for this process to happen. If Malaria is not in the salvatoy glands the mosquito cannot transmit it to humans.

The second is Aedes Egypti, which is known to transmit the Dengue virus. Unlike Malaria, Dengue can enter a mosquito and be in the salvatory glands in one day(however it can take up to 9 days).

Overall, lowering the lifespan of these mosquitos will decrease the percent chance that Malaria or Dengue will mature and reach the salvatory glands, and we know that their lifespan is somehow correlated with this "Insulin Signaling Pathway" activity. I'll try and explain.

So insulin lands on the outside of a cell and binds to a protein receptor. This protein receptor activates another protein, which activates another protein, until you reach a protein called "AKT". "AKT" is the second to last protein before transcription(beacuse the ultimate goal of this is to respond to the environmental change of the cell being in the presence of insulin). AKT promotes ISP activity, which promotes better reproductive capabilities and shorter lifespan, which is exactly what we want. At this point I am not sure why ISP activity does this, but it does. Another protein, PTEN, inhibits ISP activity. So there are two ways we can get what we want. We can kill off all the PTEN proteins and we can increase AKT protein levels.

So how do we just get rid of a protein in the body? We use viruses. So essentially a virus is double stranded RNA wrapped in a protein coat. When our immune system senses a virus, it cuts the virus into pieces(using a protein called "Dicer"), which makes the RNA single stranded. The immune system then makes a complimentary pair of this single stranded RNA and uses that track down the rest of the virus in the body. The same thing can be done to get rid of a protein. By using a single stranded RNAi (RNA interferance), we can track down the PTEN protein's RNA and destroy it. However because RNAi isn't acually written in the Genome, only the mosquito that RNAi was injected with will have less of the PTEN protein and therefore have a shorter lifespan and greater reproductive capabilities. This lets us know that IF the mosquito were to make that specific RNAi, the leves of PTEN would decrease.

We want our genetically altered mosquito to be able to reproduce faster than the already existing mosquitos. This is because if our mosquito has better fitness, then natural selection will take its course and our mosquito will out compete the wild-type mosquito. We want this to happen because our mosquito will not be able to easily transmit malaria or Dengue.

After all this was explained to me I reviewed pipeting and began learning how to isolate RNA from tissues. Despite the fact that they have an awesome machine called KaiCUBE that essentially does this for you, Anam still wants me to learn how to do it the old fashion way(which is helpful in case other labs I work in don't have an awesome machine to do it for me).

I will be working on extracting RNA from tissues by hand tomorrow and learning more about AKT and it's role in enhancing ISP throughout the week. I also expect to learn more about mosquito anatomy. I'll post again on Friday review what I did!