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A role for Kalirin-7 in nociceptive sensitization via activity-dependent modulation of spinal synapses.

Lu J, Luo C, Bali KK, Xie RG, Mains RE, Eipper BA, Kuner R - Nat Commun (2015)

Bottom Line: Synaptic plasticity is the cornerstone of processes underlying persistent nociceptive activity-induced changes in normal nociceptive sensitivity.Kalirin-7 is a multifunctional guanine-nucleotide-exchange factor (GEF) for Rho GTPases that is characterized by its localization at excitatory synapses, interactions with glutamate receptors and its ability to dynamically modulate the neuronal cytoskeleton.Here we show that spinally expressed Kalirin-7 is required for persistent nociceptive activity-dependent synaptic long-term potentiation as well as activity-dependent remodelling of synaptic spines in the spinal dorsal horn, thereby orchestrating functional and structural plasticity during the course of inflammatory pain.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, Pharmacology Institute, Medical Faculty Heidelberg, Heidelberg University, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.

ABSTRACT
Synaptic plasticity is the cornerstone of processes underlying persistent nociceptive activity-induced changes in normal nociceptive sensitivity. Kalirin-7 is a multifunctional guanine-nucleotide-exchange factor (GEF) for Rho GTPases that is characterized by its localization at excitatory synapses, interactions with glutamate receptors and its ability to dynamically modulate the neuronal cytoskeleton. Here we show that spinally expressed Kalirin-7 is required for persistent nociceptive activity-dependent synaptic long-term potentiation as well as activity-dependent remodelling of synaptic spines in the spinal dorsal horn, thereby orchestrating functional and structural plasticity during the course of inflammatory pain.

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Expression of diverse Kalirins in the spinal dorsal horn and specific knockout of Kal-7 in spinal neurons.(a) Structure of the mouse Kalrn gene and the major protein products resulting via alternative splicing. Sites targeted by the reagents used in this study are shown. (b) Western blotting with antibody against the spectrin-repeat region detects expression of Kalirin in the spinal dorsal horn (SDH), with lysates from cortex and hippocampus as positive controls. (c) Transcripts for truncated or full-length Kalirins together with full-length Kalirin-7 (Kal-7) detected in the spinal dorsal horn via RT–PCR. (d) Unilateral microinjection of recombinant adeno-associated virus (rAAV) virus expressing Cre recombinase and EGFP into the mouse lumbar spinal dorsal horn (Scale bar, 300 μm). (e) Validation of Cre-mediated Kal-7 conditional knockout in mouse spinal dorsal horn (SDH-Kal-7−/− mice) by quantitative RT–PCR, showing 50% reduction of Kal-7, but not Kal-9 or Kal-12 isoforms, in total lysate from spinal L4-L5 segments, as compared with AAV-EGFP-injected Kal-7fl/fl mice (Kal-7fl/fl-EGFP control mice) (n=3 independent experiments). *P<0.05 as compared with control group; two-way ANOVA followed by Bonferroni post-hoc test. Error bars represent s.e.m.
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f1: Expression of diverse Kalirins in the spinal dorsal horn and specific knockout of Kal-7 in spinal neurons.(a) Structure of the mouse Kalrn gene and the major protein products resulting via alternative splicing. Sites targeted by the reagents used in this study are shown. (b) Western blotting with antibody against the spectrin-repeat region detects expression of Kalirin in the spinal dorsal horn (SDH), with lysates from cortex and hippocampus as positive controls. (c) Transcripts for truncated or full-length Kalirins together with full-length Kalirin-7 (Kal-7) detected in the spinal dorsal horn via RT–PCR. (d) Unilateral microinjection of recombinant adeno-associated virus (rAAV) virus expressing Cre recombinase and EGFP into the mouse lumbar spinal dorsal horn (Scale bar, 300 μm). (e) Validation of Cre-mediated Kal-7 conditional knockout in mouse spinal dorsal horn (SDH-Kal-7−/− mice) by quantitative RT–PCR, showing 50% reduction of Kal-7, but not Kal-9 or Kal-12 isoforms, in total lysate from spinal L4-L5 segments, as compared with AAV-EGFP-injected Kal-7fl/fl mice (Kal-7fl/fl-EGFP control mice) (n=3 independent experiments). *P<0.05 as compared with control group; two-way ANOVA followed by Bonferroni post-hoc test. Error bars represent s.e.m.

Mentions: The Kalrn gene encompasses 65 exons that can be spliced in various developmentally regulated patterns to yield several different proteins, such as the full-length proteins Kalirin-7 (Kal-7), Kalirin-9 (Kal-9) and Kalirin-12 (Kal-12), as well as their N-terminally truncated versions (Δ-Kal-7; Fig. 1a)10. The longest Kalirin isoform, Kalirin-12, includes an N-terminal Sec14-like domain, homologous to yeast Sec14p; nine spectrin repeats; Rac1/RhoG-GEF domain; SH3, Src homology 3 domain; RhoA-GEF domain; a second SH3 domain followed by a C-terminal Ser/Thr protein kinase domain11. Only full-length Kal-7 and its N-terminally truncated isoform, Δ-Kal-7, possess a C-terminal PDZ-interaction motif, encoded by the first 60 nucleotides of a 3′-terminal exon included by alternative splicing; this sequence promotes interactions with PDZ domain-containing proteins such as PSD95. Direct and indirect roles for Kal-7 in both NMDAR and AMPAR localization to synapses have been demonstrated1213. Here we show that expression of specific splice variants of the Kalrn gene, particularly Kal-7, in spinal dorsal horn neurons contributes to inflammatory hypersensitivity via structure-function modulation of spinal synapses.


A role for Kalirin-7 in nociceptive sensitization via activity-dependent modulation of spinal synapses.

Lu J, Luo C, Bali KK, Xie RG, Mains RE, Eipper BA, Kuner R - Nat Commun (2015)

Expression of diverse Kalirins in the spinal dorsal horn and specific knockout of Kal-7 in spinal neurons.(a) Structure of the mouse Kalrn gene and the major protein products resulting via alternative splicing. Sites targeted by the reagents used in this study are shown. (b) Western blotting with antibody against the spectrin-repeat region detects expression of Kalirin in the spinal dorsal horn (SDH), with lysates from cortex and hippocampus as positive controls. (c) Transcripts for truncated or full-length Kalirins together with full-length Kalirin-7 (Kal-7) detected in the spinal dorsal horn via RT–PCR. (d) Unilateral microinjection of recombinant adeno-associated virus (rAAV) virus expressing Cre recombinase and EGFP into the mouse lumbar spinal dorsal horn (Scale bar, 300 μm). (e) Validation of Cre-mediated Kal-7 conditional knockout in mouse spinal dorsal horn (SDH-Kal-7−/− mice) by quantitative RT–PCR, showing 50% reduction of Kal-7, but not Kal-9 or Kal-12 isoforms, in total lysate from spinal L4-L5 segments, as compared with AAV-EGFP-injected Kal-7fl/fl mice (Kal-7fl/fl-EGFP control mice) (n=3 independent experiments). *P<0.05 as compared with control group; two-way ANOVA followed by Bonferroni post-hoc test. Error bars represent s.e.m.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4403379&req=5

f1: Expression of diverse Kalirins in the spinal dorsal horn and specific knockout of Kal-7 in spinal neurons.(a) Structure of the mouse Kalrn gene and the major protein products resulting via alternative splicing. Sites targeted by the reagents used in this study are shown. (b) Western blotting with antibody against the spectrin-repeat region detects expression of Kalirin in the spinal dorsal horn (SDH), with lysates from cortex and hippocampus as positive controls. (c) Transcripts for truncated or full-length Kalirins together with full-length Kalirin-7 (Kal-7) detected in the spinal dorsal horn via RT–PCR. (d) Unilateral microinjection of recombinant adeno-associated virus (rAAV) virus expressing Cre recombinase and EGFP into the mouse lumbar spinal dorsal horn (Scale bar, 300 μm). (e) Validation of Cre-mediated Kal-7 conditional knockout in mouse spinal dorsal horn (SDH-Kal-7−/− mice) by quantitative RT–PCR, showing 50% reduction of Kal-7, but not Kal-9 or Kal-12 isoforms, in total lysate from spinal L4-L5 segments, as compared with AAV-EGFP-injected Kal-7fl/fl mice (Kal-7fl/fl-EGFP control mice) (n=3 independent experiments). *P<0.05 as compared with control group; two-way ANOVA followed by Bonferroni post-hoc test. Error bars represent s.e.m.
Mentions: The Kalrn gene encompasses 65 exons that can be spliced in various developmentally regulated patterns to yield several different proteins, such as the full-length proteins Kalirin-7 (Kal-7), Kalirin-9 (Kal-9) and Kalirin-12 (Kal-12), as well as their N-terminally truncated versions (Δ-Kal-7; Fig. 1a)10. The longest Kalirin isoform, Kalirin-12, includes an N-terminal Sec14-like domain, homologous to yeast Sec14p; nine spectrin repeats; Rac1/RhoG-GEF domain; SH3, Src homology 3 domain; RhoA-GEF domain; a second SH3 domain followed by a C-terminal Ser/Thr protein kinase domain11. Only full-length Kal-7 and its N-terminally truncated isoform, Δ-Kal-7, possess a C-terminal PDZ-interaction motif, encoded by the first 60 nucleotides of a 3′-terminal exon included by alternative splicing; this sequence promotes interactions with PDZ domain-containing proteins such as PSD95. Direct and indirect roles for Kal-7 in both NMDAR and AMPAR localization to synapses have been demonstrated1213. Here we show that expression of specific splice variants of the Kalrn gene, particularly Kal-7, in spinal dorsal horn neurons contributes to inflammatory hypersensitivity via structure-function modulation of spinal synapses.

Bottom Line: Synaptic plasticity is the cornerstone of processes underlying persistent nociceptive activity-induced changes in normal nociceptive sensitivity.Kalirin-7 is a multifunctional guanine-nucleotide-exchange factor (GEF) for Rho GTPases that is characterized by its localization at excitatory synapses, interactions with glutamate receptors and its ability to dynamically modulate the neuronal cytoskeleton.Here we show that spinally expressed Kalirin-7 is required for persistent nociceptive activity-dependent synaptic long-term potentiation as well as activity-dependent remodelling of synaptic spines in the spinal dorsal horn, thereby orchestrating functional and structural plasticity during the course of inflammatory pain.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pharmacology, Pharmacology Institute, Medical Faculty Heidelberg, Heidelberg University, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.

ABSTRACT
Synaptic plasticity is the cornerstone of processes underlying persistent nociceptive activity-induced changes in normal nociceptive sensitivity. Kalirin-7 is a multifunctional guanine-nucleotide-exchange factor (GEF) for Rho GTPases that is characterized by its localization at excitatory synapses, interactions with glutamate receptors and its ability to dynamically modulate the neuronal cytoskeleton. Here we show that spinally expressed Kalirin-7 is required for persistent nociceptive activity-dependent synaptic long-term potentiation as well as activity-dependent remodelling of synaptic spines in the spinal dorsal horn, thereby orchestrating functional and structural plasticity during the course of inflammatory pain.

Show MeSH
Related in: MedlinePlus