A Novel Phospholipase C, PLCη2, Is a Neuron-specific Isozyme

Yoshihisa Kudo,Yoshikazu Nakamura,Yasuhiro Irino,Makoto Shimozawa,Masamichi Nakahara,Mitsuhiro Morita,Kiyoko Fukami

doi:10.1074/jbc.m503817200

Yoshihisa Kudo, Yoshikazu Nakamura + Show 5 more

Open Access

https://doi.org/10.1074/jbc.m503817200

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Abstract

Twelve phospholipase C (PLC) isozymes have been cloned so far, and they are divided into six classes, beta-, gamma-, delta-, epsilon-, zeta-, and eta-type, on the basis of structure and activation mechanisms. Here we report the identification of a novel PLC isozyme, PLC(eta)2. PLC(eta)2 is composed of conserved domains including pleckstrin homology, EF-hand, X and Y catalytic, and C2 domains and the isozyme-specific C-terminal region. PLC(eta)2 consists of 1164 amino acids with a molecular mass of 125 kDa. The PLC activity of PLC(eta)2 was more sensitive to calcium concentration than the PLC activity of the PLCdelta-type enzyme, which is thought to be the most calcium-sensitive PLC. Immunofluorescence analysis showed that PLC(eta)2 was localized predominantly to the plasma membrane at resting state via the pleckstrin homology domain. This observation was supported by Western blot analysis of cytosol and membrane fractions. In addition, expression of PLC(eta)2 was detected after birth and showed a restricted distribution in the brain; it was particularly abundant in the hippocampus, cerebral cortex, and olfactory bulb. The pattern was similar to that of the neuronal marker microtubule-associated protein 2 by Western blot. Furthermore, in situ hybridization showed positive signals for PLC(eta)2 in pyramidal cells of the hippocampus. Finally, we found that PLC(eta)2 was expressed abundantly in neuron-containing primary culture but not in astrocyte-enriched culture. These results indicate that PLC(eta)2 is a neuron-specific isozyme that may be important for the formation and/or maintenance of the neuronal network in the postnatal brain.

Highlights

Twelve phospholipase C (PLC) isozymes have been cloned so far, and they are divided into six classes, ␤, ␥, ␦, ⑀, ␨, and ␩-type, on the basis of structure and activation mechanisms
We immunoprecipitated the endogenous protein from mouse brain, and the molecular mass was compared with the molecular mass of ectopically expressed short-type (⌬N) or long-type (Full) novel PLC-like protein (Fig. 1B)
The novel protein appeared to be a member of the PLC␤ family with a long C terminus, amino acid comparison showed that it is similar to PLC␦ and has the highest similarity to PLC␩1 (Fig. 1D)

Summary

Introduction

Twelve phospholipase C (PLC) isozymes have been cloned so far, and they are divided into six classes, ␤-, ␥-, ␦-, ⑀-, ␨-, and ␩-type, on the basis of structure and activation mechanisms. Immunofluorescence analysis showed that PLC␩2 was localized predominantly to the plasma membrane at resting state via the pleckstrin homology domain This observation was supported by Western blot analysis of cytosol and membrane fractions. Expression of PLC␩2 was detected after birth and showed a restricted distribution in the brain; it was abundant in the hippocampus, cerebral cortex, and olfactory bulb. We found that PLC␩2 was expressed abundantly in neuron-containing primary culture but not in astrocyte-enriched culture These results indicate that PLC␩2 is a neuron-specific isozyme that may be important for the formation and/or maintenance of the neuronal network in the postnatal brain. PLC␩2 was abundant in primary cultured neurons but not in astrocytes These results suggest that PLC␩2 may be important for the formation and maintenance of the neuronal network in the postnatal brain

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