Mitochondrial selenium-75 uptake and regulation revealed by kinetic analysis

Area of Science:

• Biochemistry
• Environmental Science
• Insect Physiology

Background:

• Mitochondria play a crucial role in cellular metabolism and are targets for various xenobiotics.
• Selenium (Se) is an essential trace element with complex biological roles, and its uptake mechanisms are not fully understood.
• Dietary selenium levels can significantly impact cellular processes and the bioavailability of selenium compounds.

Purpose of the Study:

• To investigate the kinetics and mechanisms of selenite (Na2(75)SeO3) uptake by mitochondria from the insect C. cephalonica.
• To determine the influence of dietary selenium levels on selenite binding affinity and incorporation into mitochondrial components.
• To elucidate the specific interactions and transport pathways involved in selenite mitochondrial uptake.

Main Methods:

• Utilized radiolabeled sodium selenite (Na2(75)SeO3) to study uptake kinetics in isolated mitochondria.
• Analyzed selenite uptake across a range of concentrations and time points.
• Employed differential centrifugation to isolate mitochondrial protein fractions and assess 75Se distribution.
• Applied kinetic modeling, including Hill and Scatchard plots, to characterize binding affinities and cooperativity.
• Investigated the role of energy dependence using respiratory inhibitors and specific transport mechanisms using sulfite, sulfate, mersalyl, and NEM.

Main Results:

• Selenite uptake increased proportionally with external concentration up to 25.32 microM and linearly with time up to 15-30 min.
• Differential affinity for selenite was observed in mitochondrial protein fractions, correlating with dietary Se levels.
• Kinetic studies indicated negative cooperativity, with estimated half-saturation and high-affinity binding sites.
• Dissociation constants revealed maximal 75Se affinity in the 1.5 ppm dietary Se group.
• Energy-dependent uptake (20-30%) and involvement of the dicarboxylate port (inhibition by sulfite/sulfate) were suggested.
• Thiol-selenite interactions were confirmed by inhibition with mersalyl and NEM.

Conclusions:

• Mitochondrial selenite uptake is a complex, regulated process involving specific binding sites and energy dependence.
• Dietary selenium status significantly modulates selenite's affinity for mitochondrial binding sites and its incorporation into proteins.
• Thiol groups and potentially the dicarboxylate transporter are involved in selenite uptake, highlighting metabolic significance.
• These findings provide insights into selenium's toxicokinetics and essentiality at the mitochondrial level in insects.