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KDEL receptor 1 regulates T-cell homeostasis via PP1 that is a key phosphatase for ISR.

Kamimura D, Katsunuma K, Arima Y, Atsumi T, Jiang JJ, Bando H, Meng J, Sabharwal L, Stofkova A, Nishikawa N, Suzuki H, Ogura H, Ueda N, Tsuruoka M, Harada M, Kobayashi J, Hasegawa T, Yoshida H, Koseki H, Miura I, Wakana S, Nishida K, Kitamura H, Fukada T, Hirano T, Murakami M - Nat Commun (2015)

Bottom Line: Mechanistic analysis shows that KDELR1 directly regulates protein phosphatase 1 (PP1), a key phosphatase for ISR in naïve T cells.T-Red KDELR1 does not associate with PP1, resulting in reduced phosphatase activity against eIF2α and subsequent expression of stress responsive genes including the proapoptotic factor Bim.These results demonstrate that KDELR1 regulates naïve T-cell homeostasis by controlling ISR.

View Article: PubMed Central - PubMed

Affiliation: 1] Division of Molecular Neuroimmunology, Institute for Genetic Medicine and Graduate School of Medicine, Hokkaido University, Kita-15, Nishi-7, Kita-ku, Sapporo 060-0815, Japan [2] Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences, Graduate School of Medicine, and WPI Immunology Frontier Research Center, Osaka University, 2-2, Yamada-oka, Suita 565-0871, Japan.

ABSTRACT
KDEL receptors are responsible for retrotransporting endoplasmic reticulum (ER) chaperones from the Golgi complex to the ER. Here we describe a role for KDEL receptor 1 (KDELR1) that involves the regulation of integrated stress responses (ISR) in T cells. Designing and using an N-ethyl-N-nitrosourea (ENU)-mutant mouse line, T-Red (naïve T-cell reduced), we show that a point mutation in KDELR1 is responsible for the reduction in the number of naïve T cells in this model owing to an increase in ISR. Mechanistic analysis shows that KDELR1 directly regulates protein phosphatase 1 (PP1), a key phosphatase for ISR in naïve T cells. T-Red KDELR1 does not associate with PP1, resulting in reduced phosphatase activity against eIF2α and subsequent expression of stress responsive genes including the proapoptotic factor Bim. These results demonstrate that KDELR1 regulates naïve T-cell homeostasis by controlling ISR.

No MeSH data available.


Related in: MedlinePlus

A functional defect in KDELR1 increases stress-mediated Bim expression and apoptosis in T-Red naïve T cells.(a–d) Bim mRNA and protein levels were investigated by real-time PCR (a), western blot (b) and flow cytometry (c,d) in T-Red (T) and WT (W) mice (8–10 weeks old). N, M and B indicate naïve, memory and B cells, respectively. Expression levels of WT populations were normalized as 1 in a. Numbers in b represent the intensity ratio of Bim/Actin. Representative images from three independent experiments are shown in b,c. The mean fluorescence intensity (MFI) of intracellular staining of Bim is shown in d (n=3–4). (e) In vitro survival of naïve and memory/activated T cells in the presence or absence of IL-7. Mice between 9 and 10 weeks old were used. (f) Bim expression in naïve and memory/activated T cells by flow cytometry following retrovirus-mediated forced expression of WT KDELR1 in T-Red haematopoietic stem cells and BMT (n=4). Mice between 6 and 8 weeks were used. Data represent the mean+s.d. (a,e) or s.e.m. (d,f). P values are shown in some figures. *P<0.05; **P<0.01; ***P<0.001. NS, not significant.
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f5: A functional defect in KDELR1 increases stress-mediated Bim expression and apoptosis in T-Red naïve T cells.(a–d) Bim mRNA and protein levels were investigated by real-time PCR (a), western blot (b) and flow cytometry (c,d) in T-Red (T) and WT (W) mice (8–10 weeks old). N, M and B indicate naïve, memory and B cells, respectively. Expression levels of WT populations were normalized as 1 in a. Numbers in b represent the intensity ratio of Bim/Actin. Representative images from three independent experiments are shown in b,c. The mean fluorescence intensity (MFI) of intracellular staining of Bim is shown in d (n=3–4). (e) In vitro survival of naïve and memory/activated T cells in the presence or absence of IL-7. Mice between 9 and 10 weeks old were used. (f) Bim expression in naïve and memory/activated T cells by flow cytometry following retrovirus-mediated forced expression of WT KDELR1 in T-Red haematopoietic stem cells and BMT (n=4). Mice between 6 and 8 weeks were used. Data represent the mean+s.d. (a,e) or s.e.m. (d,f). P values are shown in some figures. *P<0.05; **P<0.01; ***P<0.001. NS, not significant.

Mentions: We next investigated why the number of naïve T cells in T-Red mice is lower than in control mice. We first considered whether naïve T cells from T-Red mice undergo higher rates of apoptosis and examined the expression levels of the proapoptotic factor Bim, a major apoptosis inducer in T cells, particularly in the periphery21222324. T-Red mice showed significantly higher expressions of Bim in their naïve T cells, but not in their memory/activated T cells or B cells when compared with controls (Fig. 5a–d). Consistent with these differences in Bim quantity between memory/activated CD4+ T cells and CD8+ T cells (Fig. 5b), it is known that memory CD8+ T cells have more and memory CD4+ T cells fewer Bim molecules than naïve T cells31 and that memory CD8+ T cells are resistant to increased expression of Bim in a manner dependent on the expression of Bcl-2 (ref. 32). Indeed, T-Red naïve T cells, but not memory/activated ones, showed lower survival rates in the presence or absence of IL-7 (Fig. 5e). We also found that Kdelr1 deficiency in naïve T cells induced more apoptosis in both CD4+ and CD8+ T cells in the presence or absence of IL-7 (Supplementary Fig. 2d). Furthermore, forced expression of the WT Kdelr1 gene decreased Bim expression in naïve T-Red T cells, but not in memory/activated ones (Fig. 5f). These results suggest that a functional defect in KDELR1 induces the expression of Bim to cause apoptosis in naïve T cells.


KDEL receptor 1 regulates T-cell homeostasis via PP1 that is a key phosphatase for ISR.

Kamimura D, Katsunuma K, Arima Y, Atsumi T, Jiang JJ, Bando H, Meng J, Sabharwal L, Stofkova A, Nishikawa N, Suzuki H, Ogura H, Ueda N, Tsuruoka M, Harada M, Kobayashi J, Hasegawa T, Yoshida H, Koseki H, Miura I, Wakana S, Nishida K, Kitamura H, Fukada T, Hirano T, Murakami M - Nat Commun (2015)

A functional defect in KDELR1 increases stress-mediated Bim expression and apoptosis in T-Red naïve T cells.(a–d) Bim mRNA and protein levels were investigated by real-time PCR (a), western blot (b) and flow cytometry (c,d) in T-Red (T) and WT (W) mice (8–10 weeks old). N, M and B indicate naïve, memory and B cells, respectively. Expression levels of WT populations were normalized as 1 in a. Numbers in b represent the intensity ratio of Bim/Actin. Representative images from three independent experiments are shown in b,c. The mean fluorescence intensity (MFI) of intracellular staining of Bim is shown in d (n=3–4). (e) In vitro survival of naïve and memory/activated T cells in the presence or absence of IL-7. Mice between 9 and 10 weeks old were used. (f) Bim expression in naïve and memory/activated T cells by flow cytometry following retrovirus-mediated forced expression of WT KDELR1 in T-Red haematopoietic stem cells and BMT (n=4). Mice between 6 and 8 weeks were used. Data represent the mean+s.d. (a,e) or s.e.m. (d,f). P values are shown in some figures. *P<0.05; **P<0.01; ***P<0.001. NS, not significant.
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Related In: Results  -  Collection

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f5: A functional defect in KDELR1 increases stress-mediated Bim expression and apoptosis in T-Red naïve T cells.(a–d) Bim mRNA and protein levels were investigated by real-time PCR (a), western blot (b) and flow cytometry (c,d) in T-Red (T) and WT (W) mice (8–10 weeks old). N, M and B indicate naïve, memory and B cells, respectively. Expression levels of WT populations were normalized as 1 in a. Numbers in b represent the intensity ratio of Bim/Actin. Representative images from three independent experiments are shown in b,c. The mean fluorescence intensity (MFI) of intracellular staining of Bim is shown in d (n=3–4). (e) In vitro survival of naïve and memory/activated T cells in the presence or absence of IL-7. Mice between 9 and 10 weeks old were used. (f) Bim expression in naïve and memory/activated T cells by flow cytometry following retrovirus-mediated forced expression of WT KDELR1 in T-Red haematopoietic stem cells and BMT (n=4). Mice between 6 and 8 weeks were used. Data represent the mean+s.d. (a,e) or s.e.m. (d,f). P values are shown in some figures. *P<0.05; **P<0.01; ***P<0.001. NS, not significant.
Mentions: We next investigated why the number of naïve T cells in T-Red mice is lower than in control mice. We first considered whether naïve T cells from T-Red mice undergo higher rates of apoptosis and examined the expression levels of the proapoptotic factor Bim, a major apoptosis inducer in T cells, particularly in the periphery21222324. T-Red mice showed significantly higher expressions of Bim in their naïve T cells, but not in their memory/activated T cells or B cells when compared with controls (Fig. 5a–d). Consistent with these differences in Bim quantity between memory/activated CD4+ T cells and CD8+ T cells (Fig. 5b), it is known that memory CD8+ T cells have more and memory CD4+ T cells fewer Bim molecules than naïve T cells31 and that memory CD8+ T cells are resistant to increased expression of Bim in a manner dependent on the expression of Bcl-2 (ref. 32). Indeed, T-Red naïve T cells, but not memory/activated ones, showed lower survival rates in the presence or absence of IL-7 (Fig. 5e). We also found that Kdelr1 deficiency in naïve T cells induced more apoptosis in both CD4+ and CD8+ T cells in the presence or absence of IL-7 (Supplementary Fig. 2d). Furthermore, forced expression of the WT Kdelr1 gene decreased Bim expression in naïve T-Red T cells, but not in memory/activated ones (Fig. 5f). These results suggest that a functional defect in KDELR1 induces the expression of Bim to cause apoptosis in naïve T cells.

Bottom Line: Mechanistic analysis shows that KDELR1 directly regulates protein phosphatase 1 (PP1), a key phosphatase for ISR in naïve T cells.T-Red KDELR1 does not associate with PP1, resulting in reduced phosphatase activity against eIF2α and subsequent expression of stress responsive genes including the proapoptotic factor Bim.These results demonstrate that KDELR1 regulates naïve T-cell homeostasis by controlling ISR.

View Article: PubMed Central - PubMed

Affiliation: 1] Division of Molecular Neuroimmunology, Institute for Genetic Medicine and Graduate School of Medicine, Hokkaido University, Kita-15, Nishi-7, Kita-ku, Sapporo 060-0815, Japan [2] Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences, Graduate School of Medicine, and WPI Immunology Frontier Research Center, Osaka University, 2-2, Yamada-oka, Suita 565-0871, Japan.

ABSTRACT
KDEL receptors are responsible for retrotransporting endoplasmic reticulum (ER) chaperones from the Golgi complex to the ER. Here we describe a role for KDEL receptor 1 (KDELR1) that involves the regulation of integrated stress responses (ISR) in T cells. Designing and using an N-ethyl-N-nitrosourea (ENU)-mutant mouse line, T-Red (naïve T-cell reduced), we show that a point mutation in KDELR1 is responsible for the reduction in the number of naïve T cells in this model owing to an increase in ISR. Mechanistic analysis shows that KDELR1 directly regulates protein phosphatase 1 (PP1), a key phosphatase for ISR in naïve T cells. T-Red KDELR1 does not associate with PP1, resulting in reduced phosphatase activity against eIF2α and subsequent expression of stress responsive genes including the proapoptotic factor Bim. These results demonstrate that KDELR1 regulates naïve T-cell homeostasis by controlling ISR.

No MeSH data available.


Related in: MedlinePlus