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Membrane palmitoylated protein 2 is a synaptic scaffold protein required for synaptic SK2-containing channel function.

Kim G, Luján R, Schwenk J, Kelley MH, Aguado C, Watanabe M, Fakler B, Maylie J, Adelman JP - Elife (2016)

Bottom Line: We have identified a novel synaptic scaffold, MPP2 (membrane palmitoylated protein 2; p55), a member of the membrane-associated guanylate kinase (MAGUK) family that interacts with SK2-containing channels.Knocking down MPP2 expression selectively abolished the SK2-containing channel contribution to synaptic responses and decreased LTP.Thus, MPP2 is a novel synaptic scaffold that is required for proper synaptic localization and function of SK2-containing channels.

View Article: PubMed Central - PubMed

Affiliation: Vollum Institute, Oregon Health and Science University, Portland, United States.

ABSTRACT
Mouse CA1 pyramidal neurons express apamin-sensitive SK2-containing channels in the post-synaptic membrane, positioned close to NMDA-type (N-methyl-D-aspartate) glutamate receptors. Activated by synaptically evoked NMDAR-dependent Ca(2+) influx, the synaptic SK2-containing channels modulate excitatory post-synaptic responses and the induction of synaptic plasticity. In addition, their activity- and protein kinase A-dependent trafficking contributes to expression of long-term potentiation (LTP). We have identified a novel synaptic scaffold, MPP2 (membrane palmitoylated protein 2; p55), a member of the membrane-associated guanylate kinase (MAGUK) family that interacts with SK2-containing channels. MPP2 and SK2 co-immunopurified from mouse brain, and co-immunoprecipitated when they were co-expressed in HEK293 cells. MPP2 is highly expressed in the post-synaptic density of dendritic spines on CA1 pyramidal neurons. Knocking down MPP2 expression selectively abolished the SK2-containing channel contribution to synaptic responses and decreased LTP. Thus, MPP2 is a novel synaptic scaffold that is required for proper synaptic localization and function of SK2-containing channels.

No MeSH data available.


MPP2 is required for synaptic SK2-containing channel function.(A) Time course of the normalized EPSP amplitude (mean ± s.e.m.) for baseline in control ACSF (Ctrl) and during wash-in of apamin (100 nM) as indicated above in MPP2 sh-transfected cells (open red symbols, n = 14) and non-fluorescent control cells (black symbols, n = 15) mice. (B) Average of 15 EPSPs taken from indicated shaded time points in aCSF (black) and 16–20 min after application of apamin (red); shaded areas are mean ± s.e.m for control non-fluorescent cells (ctrl, upper traces) and MPP2 sh-transfected cells (MPP2 sh, bottom traces). (C) Scatter plot of relative ESPS peak compared to baseline from the individual slices in panel A non-fluorescent control (ctrl, black symbols) and for MPP2 sh transfected (red symbols). Horizontal bar reflects mean response.DOI:http://dx.doi.org/10.7554/eLife.12637.010
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fig4: MPP2 is required for synaptic SK2-containing channel function.(A) Time course of the normalized EPSP amplitude (mean ± s.e.m.) for baseline in control ACSF (Ctrl) and during wash-in of apamin (100 nM) as indicated above in MPP2 sh-transfected cells (open red symbols, n = 14) and non-fluorescent control cells (black symbols, n = 15) mice. (B) Average of 15 EPSPs taken from indicated shaded time points in aCSF (black) and 16–20 min after application of apamin (red); shaded areas are mean ± s.e.m for control non-fluorescent cells (ctrl, upper traces) and MPP2 sh-transfected cells (MPP2 sh, bottom traces). (C) Scatter plot of relative ESPS peak compared to baseline from the individual slices in panel A non-fluorescent control (ctrl, black symbols) and for MPP2 sh transfected (red symbols). Horizontal bar reflects mean response.DOI:http://dx.doi.org/10.7554/eLife.12637.010

Mentions: To test whether MPP2 expression is important for synaptic SK2 channel function, two short hairpin RNAs (shRNAs) targeting the 3’ untranslated region (3’ UTR) of Mpp2 mRNA were co-expressed in area CA1 by in utero electroporation (e14-16) of a plasmid that also directed expression of the fluorescent protein, GFP. Four- to five-week-old mice were then used to prepare fresh hippocampal slices. Whole-cell current clamp recordings were made from CA1 pyramidal neurons, either transfected, as reported by GFP expression, or non-transfected control cells. Synaptic stimulations of the Schaffer collateral axons evoked EPSPs. After establishing a stable baseline, apamin (100 nM) was applied to the slices. For control cells, apamin increased EPSPs to 144.4 ± 4.3% of baseline (n = 15; p<0.0001), consistent with previous studies (Ngo-Anh et al., 2005; Lin et al., 2008; Giessel and Sabatini, 2011). In contrast, apamin did not significantly affect EPSPs from transfected CA1 pyramidal neurons (94.6 ± 3.3%; n = 14) (Figure 4A–C).10.7554/eLife.12637.010Figure 4.MPP2 is required for synaptic SK2-containing channel function.


Membrane palmitoylated protein 2 is a synaptic scaffold protein required for synaptic SK2-containing channel function.

Kim G, Luján R, Schwenk J, Kelley MH, Aguado C, Watanabe M, Fakler B, Maylie J, Adelman JP - Elife (2016)

MPP2 is required for synaptic SK2-containing channel function.(A) Time course of the normalized EPSP amplitude (mean ± s.e.m.) for baseline in control ACSF (Ctrl) and during wash-in of apamin (100 nM) as indicated above in MPP2 sh-transfected cells (open red symbols, n = 14) and non-fluorescent control cells (black symbols, n = 15) mice. (B) Average of 15 EPSPs taken from indicated shaded time points in aCSF (black) and 16–20 min after application of apamin (red); shaded areas are mean ± s.e.m for control non-fluorescent cells (ctrl, upper traces) and MPP2 sh-transfected cells (MPP2 sh, bottom traces). (C) Scatter plot of relative ESPS peak compared to baseline from the individual slices in panel A non-fluorescent control (ctrl, black symbols) and for MPP2 sh transfected (red symbols). Horizontal bar reflects mean response.DOI:http://dx.doi.org/10.7554/eLife.12637.010
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Related In: Results  -  Collection

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fig4: MPP2 is required for synaptic SK2-containing channel function.(A) Time course of the normalized EPSP amplitude (mean ± s.e.m.) for baseline in control ACSF (Ctrl) and during wash-in of apamin (100 nM) as indicated above in MPP2 sh-transfected cells (open red symbols, n = 14) and non-fluorescent control cells (black symbols, n = 15) mice. (B) Average of 15 EPSPs taken from indicated shaded time points in aCSF (black) and 16–20 min after application of apamin (red); shaded areas are mean ± s.e.m for control non-fluorescent cells (ctrl, upper traces) and MPP2 sh-transfected cells (MPP2 sh, bottom traces). (C) Scatter plot of relative ESPS peak compared to baseline from the individual slices in panel A non-fluorescent control (ctrl, black symbols) and for MPP2 sh transfected (red symbols). Horizontal bar reflects mean response.DOI:http://dx.doi.org/10.7554/eLife.12637.010
Mentions: To test whether MPP2 expression is important for synaptic SK2 channel function, two short hairpin RNAs (shRNAs) targeting the 3’ untranslated region (3’ UTR) of Mpp2 mRNA were co-expressed in area CA1 by in utero electroporation (e14-16) of a plasmid that also directed expression of the fluorescent protein, GFP. Four- to five-week-old mice were then used to prepare fresh hippocampal slices. Whole-cell current clamp recordings were made from CA1 pyramidal neurons, either transfected, as reported by GFP expression, or non-transfected control cells. Synaptic stimulations of the Schaffer collateral axons evoked EPSPs. After establishing a stable baseline, apamin (100 nM) was applied to the slices. For control cells, apamin increased EPSPs to 144.4 ± 4.3% of baseline (n = 15; p<0.0001), consistent with previous studies (Ngo-Anh et al., 2005; Lin et al., 2008; Giessel and Sabatini, 2011). In contrast, apamin did not significantly affect EPSPs from transfected CA1 pyramidal neurons (94.6 ± 3.3%; n = 14) (Figure 4A–C).10.7554/eLife.12637.010Figure 4.MPP2 is required for synaptic SK2-containing channel function.

Bottom Line: We have identified a novel synaptic scaffold, MPP2 (membrane palmitoylated protein 2; p55), a member of the membrane-associated guanylate kinase (MAGUK) family that interacts with SK2-containing channels.Knocking down MPP2 expression selectively abolished the SK2-containing channel contribution to synaptic responses and decreased LTP.Thus, MPP2 is a novel synaptic scaffold that is required for proper synaptic localization and function of SK2-containing channels.

View Article: PubMed Central - PubMed

Affiliation: Vollum Institute, Oregon Health and Science University, Portland, United States.

ABSTRACT
Mouse CA1 pyramidal neurons express apamin-sensitive SK2-containing channels in the post-synaptic membrane, positioned close to NMDA-type (N-methyl-D-aspartate) glutamate receptors. Activated by synaptically evoked NMDAR-dependent Ca(2+) influx, the synaptic SK2-containing channels modulate excitatory post-synaptic responses and the induction of synaptic plasticity. In addition, their activity- and protein kinase A-dependent trafficking contributes to expression of long-term potentiation (LTP). We have identified a novel synaptic scaffold, MPP2 (membrane palmitoylated protein 2; p55), a member of the membrane-associated guanylate kinase (MAGUK) family that interacts with SK2-containing channels. MPP2 and SK2 co-immunopurified from mouse brain, and co-immunoprecipitated when they were co-expressed in HEK293 cells. MPP2 is highly expressed in the post-synaptic density of dendritic spines on CA1 pyramidal neurons. Knocking down MPP2 expression selectively abolished the SK2-containing channel contribution to synaptic responses and decreased LTP. Thus, MPP2 is a novel synaptic scaffold that is required for proper synaptic localization and function of SK2-containing channels.

No MeSH data available.