A highly expressing Tet-inducible cell line recapitulates in situ developmental changes in prestin's Boltzmann characteristics and reveals early maturational events.
Prestin is the motor protein within the lateral membrane of outer hair cells (OHCs), and it is required for mammalian cochlear amplification. Expression of prestin precedes the onset of hearing in mice, and it has been suggested that prestin undergoes a functional maturation within the membrane coincident with the onset of hearing. We have developed a tetracycline-inducible prestin-expressing cell line that we have used to model prestin's functional maturation. We used prestin's voltage-dependent nonlinear charge movement (or nonlinear capacitance) as a test of function and correlated it to biochemical measures of prestin expressed on the cell surface. An initial stage of slow growth in charge density is accompanied by a rapid increase in our estimate of charge carried by an individual motor. A rapid growth in charge density follows and strongly correlates with an increasing ratio between an apparently larger and smaller monomer, suggesting that the latter exerts a dominant-negative effect on function. Finally, there is a gradual depolarizing shift in the voltage of peak capacitance, similar to that observed in developing OHCs. This inducible system offers many opportunities for detailed studies of prestin.
Bian
Shumin
S
Department of Neurology, Yale School of Medicine, New Haven, CT 06520, USA.
Koo
Bon W
BW
Kelleher
Stephen
S
Santos-Sacchi
Joseph
J
Navaratnam
Dhasakumar S
DS
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003051:Cochlea / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism*; D025802:Electric Capacitance; D018072:Hair Cells, Auditory, Outer / Q000166:cytology / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D015640:Ion Channel Gating / Q000502:physiology*; D051379:Mice; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D020409:Molecular Motor Proteins; C498720:Pres protein, mouse; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins