Functional characterization of slac1 knockout in Gossypium hirsutum using physiological and metabolic measurements

Cotton (Gossypium hirsutum) is an agronomically important crop that requires substantial water. Cotton is historically grown in areas with low precipitation, where drought is likely to become more severe. Stomata are microscopic pores on the leaf surface that regulate gas exchange and transpiration, making their function critical for balancing photosynthesis with water conservation. The SLAC1 (Slow Anion Channel-Associated 1) protein is a key component of the stomatal closure mechanism under environmental stress and is highly conserved across plant lineages. However, SLAC1 is not essential for plant survival, indicating the presence of alternative pathways for mitigating water stress. This study investigates how slac1 CRISPR knockout cotton plants respond to drought stress at both physiological and metabolic levels in the absence of functional stomatal closure. Water-limiting experiments were conducted using LI-6800 and FluorPen instrumentation to assess gas exchange and photosynthetic efficiency. Concurrently, metabolite profiling focused on starch and antioxidant pathways, guided by differential gene expression analysis, which revealed upregulation of genes involved in starch biosynthesis and antioxidant activity in slac1 mutants relative to wild-type plants. Physiological measurements confirmed that slac1 mutants exhibit consistently higher stomatal conductance across all levels of soil water content, consistent with impaired closure. Metabolomic assays further revealed significantly elevated antioxidant activity in slac1 mutants, particularly under drought stress (p = 0.00052), while no significant difference in starch accumulation was observed between genotypes (p = 0.9098). These findings suggest that slac1 cotton plants may compensate for impaired stomatal regulation by enhancing antioxidant defenses, highlighting an alternative stress response pathway. This insight could inform strategies for engineering drought-resilient cotton cultivars that can maintain productivity under water-limited conditions.

I am thankful to be welcomed back to the Boyce Thompson Institute (BTI) this summer to continue expanding my knowledge of plant biology research. My early experience at BTI began in high school, when I was uncertain about my future, but being able to return and continue exploring drought stress research has been invaluable. I appreciate my mentor and the Nelson Lab for welcoming me, and working on an NSF-funded CROPPS project has been a meaningful milestone.

Focusing my research on how drought stress affects stomatal regulation in cotton through the Slow Anion Channel 1 (SLAC1), I investigated a critical mechanism for plant water conservation. Direct engagement with plant stress physiology enabled me to expand my expertise and enter new areas, such as bioinformatics. Throughout the internship, I gained experience with a variety of scientific techniques and instruments. For example, I worked with the LI-6800, LI-600, and FluorPen, performing DPPH assays to evaluate antioxidant capacity and quantifying starch to assess metabolic changes. These experiences contributed to my overall understanding and personal growth during the program.

I am grateful to have had the opportunity to participate in this internship. Not only did it provide valuable insights into my career path, but it also gave us access to resources that keep us informed about the world of plant science. My understanding of agricultural resilience expanded, and I was introduced to the possibilities of graduate research. Although there is still much to learn about plants, being part of the REU program at BTI allowed me to form lasting professional connections and gain inspiration for my future in science, an incredible opportunity.
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