“Characterization of a Putative Cutinase from Tomato Fruits”
The cuticle, an extracellular layer of fatty acids which coats the outer aerial surface of all land plants. By mass, the cuticle is predominantly composed of cutin, a matrix of polyhydroxylated fatty acids. To date, many of the genes responsible for the formation of cutin that have been identified are biosynthetic in nature leading to little understanding of the mechanism of cuticle assembly and disassembly in the extracellular space. Previously, a protein within the GDSL esterase/lipase superfamily of proteins has been reported to degrade polymeric cutin in Arabidopsis and was deemed cuticle destructing factor (AtCDEF). However, no phenotype was identified in lines with reduced CDEF1 expression and no physiological relevance had been determined, due in large part to the difficulty of analyzing the cuticle of Arabidopsis. To this end, the thick, astomatous cuticle of tomato fruits offer an ideal system for upstanding cuticle biology and has led to the identification of another GDSL protein which is responsible for the polymerization of the majority of cutin in tomato fruit. By using available amino acid sequences and transcript expression data, an ortholog of AtCDEF was identified which is expressed in the outer epidermis of tomato fruits, SlCDEF, and may be involved in the degradation of cutin. The goals of my project have been twofold: to confirm that SlCDEF is able to depolymerize polymeric cutin in vitro using mass spectrometry and to demonstrate an in planta function of SlCDEF in tomato fruits using CRISPR technology.
Spending the summer in the BTI REU program has allowed me to gain experience with new research techniques while trying to answer a fascinating research question. My project allowed me to work with biochemical, genetic, and molecular techniques I had briefly experienced in classes, but never used in a research setting. In addition, I was able to learn more about the wide variety of interesting research occurring at BTI, which has allowed me to think more about my plans for research in graduate school. Experiencing research at BTI as well as speaking with faculty and graduate students allowed me to think more of my own research goals, as well as the skills that would be critical to accomplishing them. Ultimately, I feel that spending the summer at BTI has broadened my research skills and better prepared me for applying to graduate school next year.