Amyloidogenic lysozymes accumulate in the endoplasmic reticulum accompanied by the augmentation of ER stress signals

Yoshiki Kamada, Takahiro Kusakabe, Yasushi Sugimoto

研究成果: ジャーナルへの寄稿記事

4 引用 (Scopus)

抄録

Background: Naturally occurring single mutants, I56T, F57I, W64R and D67H of lysozyme in human, have been known to form abnormal protein aggregates (amyloid fibrils) and to accumulate in several organs, including the liver, spleen and kidney, resulting in familial systemic amyloidosis. These human pathogenic lysozyme variants are considered to raise subtle conformational changes compared to the wild type. Methods: Here we examined the effects of the aberrant mutant lysozymes I56T, F57I, W64R and D67H, each of which possesses a point mutation in its molecule, on a cultured human cell line, HEK293, in which the genes were individually integrated and overexpressed. Results: Western blot analyses showed lesser amounts of these variant proteins in the medium compared to the wild type, but they were abundant in the cell pellets, indicating that the modified lysozyme proteins were scarcely secreted into the medium but were retained in the cells. Immunocytochemistry revealed that these proteins resided in restricted regions which were stained by an endoplasmic reticulum (ER) marker. Moreover, the overexpression of the mutant lysozymes were accompanied by marked increases in XBP-1s and GRP78/BiP, which are downstream agents of the IRE1α signaling pathway responding to the unfolded protein response (UPR) upon ER stress. RNAi for the mutant lysozymes' expression greatly suppressed the increases of these agents. Conclusions and General significance: Our results suggest that the accumulation of pathogenic lysozymes in the ER caused ER stress and the UPR response mainly via the IRE1α pathway.

元の言語英語
ページ(範囲)1107-1119
ページ数13
ジャーナルBiochimica et Biophysica Acta - General Subjects
1850
発行部数6
DOI
出版物ステータス出版済み - 1 1 2015

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Endoplasmic Reticulum Stress
Muramidase
Endoplasmic Reticulum
Unfolded Protein Response
Proteins
Familial Amyloidosis
Heat-Shock Proteins
RNA Interference
Point Mutation
Amyloid
Liver
Cultured Cells
Spleen
Genes
Western Blotting
Immunohistochemistry
Cells
Kidney
Cell Line
Molecules

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

これを引用

Amyloidogenic lysozymes accumulate in the endoplasmic reticulum accompanied by the augmentation of ER stress signals. / Kamada, Yoshiki; Kusakabe, Takahiro; Sugimoto, Yasushi.

:: Biochimica et Biophysica Acta - General Subjects, 巻 1850, 番号 6, 01.01.2015, p. 1107-1119.

研究成果: ジャーナルへの寄稿記事

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abstract = "Background: Naturally occurring single mutants, I56T, F57I, W64R and D67H of lysozyme in human, have been known to form abnormal protein aggregates (amyloid fibrils) and to accumulate in several organs, including the liver, spleen and kidney, resulting in familial systemic amyloidosis. These human pathogenic lysozyme variants are considered to raise subtle conformational changes compared to the wild type. Methods: Here we examined the effects of the aberrant mutant lysozymes I56T, F57I, W64R and D67H, each of which possesses a point mutation in its molecule, on a cultured human cell line, HEK293, in which the genes were individually integrated and overexpressed. Results: Western blot analyses showed lesser amounts of these variant proteins in the medium compared to the wild type, but they were abundant in the cell pellets, indicating that the modified lysozyme proteins were scarcely secreted into the medium but were retained in the cells. Immunocytochemistry revealed that these proteins resided in restricted regions which were stained by an endoplasmic reticulum (ER) marker. Moreover, the overexpression of the mutant lysozymes were accompanied by marked increases in XBP-1s and GRP78/BiP, which are downstream agents of the IRE1α signaling pathway responding to the unfolded protein response (UPR) upon ER stress. RNAi for the mutant lysozymes' expression greatly suppressed the increases of these agents. Conclusions and General significance: Our results suggest that the accumulation of pathogenic lysozymes in the ER caused ER stress and the UPR response mainly via the IRE1α pathway.",
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N2 - Background: Naturally occurring single mutants, I56T, F57I, W64R and D67H of lysozyme in human, have been known to form abnormal protein aggregates (amyloid fibrils) and to accumulate in several organs, including the liver, spleen and kidney, resulting in familial systemic amyloidosis. These human pathogenic lysozyme variants are considered to raise subtle conformational changes compared to the wild type. Methods: Here we examined the effects of the aberrant mutant lysozymes I56T, F57I, W64R and D67H, each of which possesses a point mutation in its molecule, on a cultured human cell line, HEK293, in which the genes were individually integrated and overexpressed. Results: Western blot analyses showed lesser amounts of these variant proteins in the medium compared to the wild type, but they were abundant in the cell pellets, indicating that the modified lysozyme proteins were scarcely secreted into the medium but were retained in the cells. Immunocytochemistry revealed that these proteins resided in restricted regions which were stained by an endoplasmic reticulum (ER) marker. Moreover, the overexpression of the mutant lysozymes were accompanied by marked increases in XBP-1s and GRP78/BiP, which are downstream agents of the IRE1α signaling pathway responding to the unfolded protein response (UPR) upon ER stress. RNAi for the mutant lysozymes' expression greatly suppressed the increases of these agents. Conclusions and General significance: Our results suggest that the accumulation of pathogenic lysozymes in the ER caused ER stress and the UPR response mainly via the IRE1α pathway.

AB - Background: Naturally occurring single mutants, I56T, F57I, W64R and D67H of lysozyme in human, have been known to form abnormal protein aggregates (amyloid fibrils) and to accumulate in several organs, including the liver, spleen and kidney, resulting in familial systemic amyloidosis. These human pathogenic lysozyme variants are considered to raise subtle conformational changes compared to the wild type. Methods: Here we examined the effects of the aberrant mutant lysozymes I56T, F57I, W64R and D67H, each of which possesses a point mutation in its molecule, on a cultured human cell line, HEK293, in which the genes were individually integrated and overexpressed. Results: Western blot analyses showed lesser amounts of these variant proteins in the medium compared to the wild type, but they were abundant in the cell pellets, indicating that the modified lysozyme proteins were scarcely secreted into the medium but were retained in the cells. Immunocytochemistry revealed that these proteins resided in restricted regions which were stained by an endoplasmic reticulum (ER) marker. Moreover, the overexpression of the mutant lysozymes were accompanied by marked increases in XBP-1s and GRP78/BiP, which are downstream agents of the IRE1α signaling pathway responding to the unfolded protein response (UPR) upon ER stress. RNAi for the mutant lysozymes' expression greatly suppressed the increases of these agents. Conclusions and General significance: Our results suggest that the accumulation of pathogenic lysozymes in the ER caused ER stress and the UPR response mainly via the IRE1α pathway.

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