Ca2+ binding to Esyt is required to modulate membrane contact site density in Drosophila photoreceptors

Abstract

Membrane Contact Sites (MCS) between the plasma membrane (PM) and endoplasmic reticulum (ER) have been shown to regulate Ca2+ influx into animal cells. However, the mechanisms by which cells modulate ER-PM MCS density is not understood and the role of Ca2+, if any, in regulating this process is not known. We report that in Drosophila photoreceptors, MCS density is dependent on the activity of the Ca2+ permeable channels-TRP and TRPL. This regulation of MCS density by Ca2+ is mediated by extended synaptotagmin (dEsyt), a protein localised to ER-PM MCS in photoreceptors and previously shown to regulate MCS density. We find that the Ca2+ binding activity of dEsyt is required for its functional activity in vivo. dEsytCaBM, a Ca2+ non-binding mutant of dEsyt is unable to modulate MCS structure in a manner equivalent to its wild type counterpart. Further, when reconstituted in dEsyt null photoreceptors, in contrast to wild type dEsyt, dEsytCaBM is unable to rescue ER-PM MCS density and other key phenotypes. Finally, when expressed in wild type photoreceptors, dEsytCaBM phenocopies loss of dEsyt function. Taken together, our data supports a role for the Ca2+ binding activity of dEsyt in regulating the ER-PM MCS density in photoreceptors thus tuning signal transduction in response to Ca2+ influx triggered by ambient illumination. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=196 SRC="FIGDIR/small/588361v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@f8faacorg.highwire.dtl.DTLVardef@114009org.highwire.dtl.DTLVardef@8337f0org.highwire.dtl.DTLVardef@cd7c2f_HPS_FORMAT_FIGEXP M_FIG C_FIG