Katherine Koning
Year: 2021
Faculty Advisor: Miguel Pineros

“Regulation of Apple Acidity: Is it Protein Turnover of ALMT Transporter?”

Project Summary:

The aluminum-activated malate transporter (ALMT) family of ion channels plays an essential role in plant mineral nutrition processes, fruit quality, and responses to abiotic stress. Transport facilitated by membrane transport proteins, including those in the ALMT family, is regulated at the transcriptional and post-translational levels. In apples, the regulation of the apple ALMT9 transporter influences its function, and consequently, the levels of the fruit’s acidity. Previous work in wheat and apple has shown that truncation of the C-terminus of the ALMT transporter protein abolished its function and decreased its expression, as seen by a reduction in transient transformation efficiency. This project aims to determine whether these observations are due to differences in ALMT regulation resulting from differences in protein turnover.

Using tandem fluorescent timer (tFT) chimeras transiently expressed in N. benthamiana, we examined protein turnover of two apple ALMT9 alleles associated with fruit acidity, differing in their functional and expression properties. Given the different maturation rates of the fluorophores, their ratios could be used to compare protein lifetime. The relative lifetimes of two allelic variants, one full length and one with a truncated C-terminus, were determined using the ratio between the intensities of the fluorophores in the tFT. The combination of the turnover data inferred from the tFTs with previous functional observations will allow us to discern if these structural differences underlined by the C-terminal truncation alter the transport characteristics of these proteins or whether these modifications have a larger effect on the relative lifetime of these proteins. Understanding the molecular mechanism by which this transporter is regulated will result in new molecular targets to adjust apple acidity for commercial purposes.

My Experience:

As a member of the Piñeros lab this summer, I gained confidence working collaboratively and independently on my project. I was welcomed and encouraged by my mentor, PI, and other members of the lab.

My experiences both within the lab and seminar portions of the internship gave me clarity about my research interests and graduate school application process. I gained insight into life as a graduate student and my goals for my graduate school experience. The training I received in bioinformatics and confocal microscopy will be extremely valuable as I continue pursuing a career in research.