Mpp4 at the Photoreceptor Synaptic Terminals: Anchoring Plasma Membrane Calcium ATPases and Modulating Calcium Homeostasis

Abstract

Purpose:: Membrane palmitoylated protein 4 (Mpp4) is a photoreceptor-specific protein belonging to the membrane-associated guanylate kinase family, with unknown biological functions. We carried out genetic, cell biological and biochemical studies to define the function of Mpp4 in photoreceptor cells.

Methods:: The Mpp4 gene was disrupted by gene targeting in mice. Phenotype analysis and protein functional studies utilized light and electron microscopy, immunofluorescence, Western blotting, immunoprecipitation, yeast two-hybrid protein interaction analysis, electroretinography (ERG), and single cell recordings.

Results:: Mpp4 localizes specifically to the synaptic terminals of both rod and cone photoreceptors. Plasma membrane Ca²⁺ ATPases (PMCAs), the Ca²⁺ extrusion pumps, interact with an Mpp4-dependent presynaptic membrane protein complex that includes Veli3 and PSD95. In mice lacking Mpp4, PMCAs are lost from the photoreceptor pre-synaptic membranes. Synaptic ribbons are enlarged. Sarcoplasmic-endoplasmic reticulum Ca²⁺ ATPase, type 2, localizing close to the ribbons, is increased. The distribution of InsP₃ receptor, type 2 shifts from the synaptic terminal to the basal outer segment. Although single cell recordings show largely intact phototransduction, delays in the timing and reduced amplitudes of ERG b-waves and oscillatory potentials indicate that synaptic transmission to second-order retinal neurons is abnormal.

Conclusions:: Loss of Mpp4 disrupts a major Ca²⁺ extrusion mechanism at the presynaptic membranes, with ensuing adaptive responses by the photoreceptor cell to restore Ca²⁺ homeostasis. We conclude that Mpp4 affects Ca²⁺ homeostasis and modulates synaptic transmission in photoreceptors by organizing a presynaptic membrane protein complex that includes PMCAs.

Keywords: photoreceptors • transgenics/knock-outs • calcium