Research Tribute

Looking into a Cell

Vickie J. LaMorte, Ph.D., is an Associate Professor at the Beckman Laser Institute. She has been at BLI for the past eight years developing ways to utilize optical techniques to wxamine the in vivo function of nuclear proteins. To do this , she is bridgeing traditional molecular biology techniques with biophotonics to develop new approaches to monitor a protein's function inside the cell. by utilizing advanced optical mircoscopy techniques, she is able "to peer into the cell" and monitor what a particular protein is doing.

Through funding from the American Heart Association, she was able to monitor proteins that play a very important role in cholesterol metabolism. Her findings were recently published in the April issue of Molecular and Cellular Biology entitled "Spatial Distribution and Function of Sterol Reglatory Element-binding Protein 1a and 2 Homo- and Heterodimers by In Vivo Two-Photon Imaging Spectroscopy and Flurorescence Reasonance Energy Transfer."

Fluorescence resonance energy transfer detection of SREBP-1a
and SREBP-2 heterodimerization in vivo: a two-photon
fuorescence image of a single cell expressing
SREBP-1aCFP/SREBP-2YFP in a foci pattern.

Cholesterol is a vital molecule for an array of biological processes as both a structural building block and a precursor to a host of signaling ligands. As imprtant as cholesterol is to the vitality of the organism, it can be deleterious to its survival since there is no way to recover engery and carbon from this molecule. this results in a clinically well-described accumulation and deposition in the blood vessels where the blood flow is impeded. Key players in cholesterol regulation are the members of a family of transcription factors known as the Sterol Reglatory Binding Proteins (SREBPs). Transcription factors are proteins that bind to DNA and acty as molecular switches to turn genes on or off. The celluelar redundancy of this family of SREBP proteins is under investigation. What regulates these proteins to allow them to have differential roles in the cell and, ultimately, control over when certain genes are switched on or off is key to developing new drugs.

Understanding the molecular dynamics of these SREBP porteins and how they function in the highly organized enviornment of the nucleus may provide clues to deciphering the cell's requirement for expressing seemingly similar proteins. By utilizing a fluorescent protein found in jellyfish, one can tag these different SREBP proteins and follow their movement within a living cell. this is done by engineering the DNA sequence of the jellyfish protein into the DNA sequence of the protein one wants to track. In essence, a fusion protein is created.

All of these advanced studies require recombinant DNA, and in order to isolate and re-engineer it, one needs the technology that Dr. Beckman developed. Without a pH meter, one would not be able to mix solutions that are compatible with the inside of the cell. A spectrophotometer is needed to know how much DNA or proteins are are in the tube. And without a centrifuge, one would not be able to isolate the DNA from the bacterial cells that one commandeers to make the recombiant DNA.

From here, Dr. LaMorte id able to introduce the DNA into the cells with a very thin glass needle. After that, it is up to the cell to read the DNA and make the protein. Once the cell is expressing the glowing protein, Dr. LaMorte can utilize advanced optical tools to monitor this protein in the cell. She is able to know not only where the protein is going but with whom it is talking to along the way. Dr. LaMorte's observations of these proteins indicate a dynamic association between a specific member of this family, SREBP-2 and the PML-containing nuclear body, unlike its sister portein, SREBP-1, which does not appear to associate with this nuclear body. It was also demostrated for the first time which proteins partner or "talk to each other" and when.

It is sometimes forgotten that basic instrumentation is the first step needed to advanc studies like Dr. LaMorte's and her lab. grateful scientists everywhere offer special thank you to Dr. Beckamn for inventing the basic technological building blocks that are the keystones to research today.