Title

Cloning and Expression of Metallothionein Genes from S. Cerevisiae and A. Fumigatus

School Name

South Carolina Governor's School for Science and Mathematics

Grade Level

12th Grade

Presentation Topic

Cell and Molecular Biology

Presentation Type

Mentored

Abstract

Transition metals, such as copper, are key components of biological systems. Because of their special properties, they are required for the function of proteins that perform essential cellular processes. However, having excess metals in the cell can also be toxic to cell growth. Metallothioneins (MT) are proteins that can help relieve this toxicity by binding to excess metals in the cell. The metallothioneins being studied are copper metallothioneins: Cup1 and Crs5 from Saccharomyces cerevisiae, and CmtA from Aspergillus fumigatus. The genes for these proteins are expressed only when their promoters are activated. To study the impact of high gene expression on cell survival, the constitutive TEF1 promoter is used. The TEF1 promoter can be used to constantly overexpress the genes that correlate to metallothionein production. The goal was to construct a plasmid that uses the TEF1 promoter to drive MT gene expression so that it can be used for future growth experiments. Plasmids with the MT genes, CUP1, CRS5, and CMTA, were created by genetic engineering including steps such as subcloning, polymerase chain reactions, the use of restriction enzyme digestion, and Gibson Assembly. The resulting plasmids included both the TEF1 promoter and a metallothionein gene to be studies. An ultimate goal that can be explored in further research is to determine how high expression of these MT proteins impacts cells survival under high and low Cu conditions, which might occur during infection.

Location

HSS 203

Start Date

4-2-2022 12:00 PM

Presentation Format

Oral Only

Group Project

Yes

COinS
 
Apr 2nd, 12:00 PM

Cloning and Expression of Metallothionein Genes from S. Cerevisiae and A. Fumigatus

HSS 203

Transition metals, such as copper, are key components of biological systems. Because of their special properties, they are required for the function of proteins that perform essential cellular processes. However, having excess metals in the cell can also be toxic to cell growth. Metallothioneins (MT) are proteins that can help relieve this toxicity by binding to excess metals in the cell. The metallothioneins being studied are copper metallothioneins: Cup1 and Crs5 from Saccharomyces cerevisiae, and CmtA from Aspergillus fumigatus. The genes for these proteins are expressed only when their promoters are activated. To study the impact of high gene expression on cell survival, the constitutive TEF1 promoter is used. The TEF1 promoter can be used to constantly overexpress the genes that correlate to metallothionein production. The goal was to construct a plasmid that uses the TEF1 promoter to drive MT gene expression so that it can be used for future growth experiments. Plasmids with the MT genes, CUP1, CRS5, and CMTA, were created by genetic engineering including steps such as subcloning, polymerase chain reactions, the use of restriction enzyme digestion, and Gibson Assembly. The resulting plasmids included both the TEF1 promoter and a metallothionein gene to be studies. An ultimate goal that can be explored in further research is to determine how high expression of these MT proteins impacts cells survival under high and low Cu conditions, which might occur during infection.