Every 30 minutes a child is born with a mitochondrial disease. By the time they are 10 years old, they will experience symptoms that include seizures, weakness and an inability to fight infection. This is a result of the mitochondria failing to convert oxygen and nutrients into ATP energy that the body needs to function. There is no known cure for a mitochondrial disease because the biology of this energy-producing organelle is not completely known.
In the semester of 2016 in Molecular Genetics Research with Dr. Ed Turk, students experimented with yeast to identify proteins that enter the mitochondria. This knowledge could further the understanding of mitochondrial biology, and thus lead to cures for mitochondrial diseases.
Students used yeast cells as a human cell simulation because the cellular make-up of yeast is very similar to that of a human cell. Yeast are single-celled microorganisms; and a type of fungi. The yeast that was used, Saccharomyces cerevisae, is the same type of yeast that is used to make bread, so it is not harmful to work with. Yeast and human cells are both Eukaryotic, meaning that through experimenting with yeast, students can learn about the function of the human cell.
Eukaryotic cells are complex and contain membrane enclosed organelles such as a nucleus containing DNA bound together with proteins. For comparison, bacteria are Prokaryotic cells that are much smaller than Eukaryotic cells. They contain mostly cytoplasm with DNA not surrounded by a membrane. In addition to being easy to manipulate, yeast cells resemble human cells because they both contain the organelle, mitochondria.
Mitochondria is commonly known as the “powerhouse of the cell.” It is responsible for producing ATP (energy) that the cell needs to function. It is a double-membraned organelle that carries out cellular respiration. It consists of three main parts: Outer membrane, Inner Membrane, and Mitochondrial Matrix.
To understand how the mitochondria can be controlled by proteins, students must first understand the biology and function of the protein. Proteins are little machines made of organic materials that carry out specific functions to keep our bodies strong and healthy. They are long chains of amino acids (monomers) that are bonded together to create polypeptides. Proteins carry the translated information from the RNA which was transcribed from the genetic information of DNA. Proteins are the product of the final step of the Central Dogma.
The Central Dogma is the “term” used to explain how information is carried from DNA to physical traits like eye color, for example (because they are controlled by proteins). DNA is the genetic information in the chromosomes (which contain a double helix of nucleotides). RNA is a single stranded nucleic acid that is transcribed from base pairing nucleotides from the DNA. Finally, the RNA is translated into amino acids that form the proteins. The information held within the genetic makeup of the DNA essentially determines the proteins that are formed, which is why the only way to manipulate proteins is through manipulating the DNA (which can be done through plasmid manipulation).
Plasmids are small circular strands of DNA that replicate independently of chromosomal DNA at origins of replication. Students worked with plasmids because they are very easy to manipulate and the yeast cells can absorb them. By manipulating the gene (plasmid), students manipulate the protein they are interested in. In summary, DNA that stores the genetic information (in the plasmid) is altered in order to change the RNA in order to change the protein.
The main purpose of the experiment was to determine if the NGL1 protein enters the mitochondria. The NGL1 gene was inserted into a plasmid and fused with the ARG8 gene. The protein I tested (NGL1) entered the mitochondria and dragged along the -ARG8 protein with it. Since the yeast cells grew, that means that the protein we tested must have dragged in ARG8 with it because it contained a signal that told it to enter the mitochondria; therefore producing arginine in the mitochondria.
Finally, students know what they know simply because the yeast cells visibly grew. One can observe that yeast cells on a plate are growing. Since students cannot see the microscopic reactions taking place, they must rely on broader observations, such as growth. Therefore, they know the protein must have entered the mitochondria (on the -ARG plate) because it grew. This all means that the gene (NGL1) coded for a protein that dragged –ARG8 into the mitochondria. This seems promising considering this protein was able to enter the mitochondria. However, some yeast cells showed no growth for certain proteins. In Molecular Genetics Research, other peers experiment with different proteins to conduct multiple trials. However, further research needs to be conducted to verify these results and understanding of mitochondrial biology.