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Calreticulin is a negative regulator of bronchial smooth muscle cell proliferation.

Miglino N, Roth M, Lardinois D, Tamm M, Borger P - J Allergy (Cairo) (2012)

Bottom Line: In the presence of 5% FBS, the p42/p30 ratio significantly decreased (n = 3, P < 0.05) and coincided with BSM cell proliferation.High levels of calreticulin were associated with a decreased p42/p30 isoform ratio.Modulation of calreticulin levels may provide a novel target to reduce BSM remodeling.

View Article: PubMed Central - PubMed

Affiliation: Departments of Biomedicine, Pulmonary Cell Research, and Thorax Surgery, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland.

ABSTRACT
Background. Calreticulin controls the C/EBPαp42/p30 at the translational level trough a cis-regulatory CNG rich loop in the CEBPA mRNA. We determined the effects of steroids and long-acting beta-agonists on the p42/p30 ratio and on calreticulin expression in primary human bronchial smooth muscle (BSM) cells. Methods. The effects of budesonide (10(-8) M) and formoterol (10(-8) M) were studied in BSM cells pre-treated with siRNA targeting calreticulin. The expression of C/EBPα and calreticulin was determined by immuno-blotting. Automated cell counts were performed to measure proliferation. Results. All tested BSM cell lines (n = 5) expressed C/EBPα and calreticulin. In the presence of 5% FBS, the p42/p30 ratio significantly decreased (n = 3, P < 0.05) and coincided with BSM cell proliferation. High levels of calreticulin were associated with a decreased p42/p30 isoform ratio. FBS induced the expression of calreticulin (n = 3, P < 0.05), which was further increased by formoterol. siRNA targeting calreticulin increased the p42/p30 ratio in non-stimulated BSM cells and significantly inhibited the proliferation of PDGF-BB-stimulated BSM cells (n = 5, P < 0.05). Neither budesonide nor formoterol restored the p42 isoform expression. Conclusions. Our data show calreticulin is a negative regulator of C/EBPα protein expression in BSM cells. Modulation of calreticulin levels may provide a novel target to reduce BSM remodeling.

No MeSH data available.


(a) Simplified scheme of the CEBPA mRNA. Due to alternative translation start sites (A, B, and C), full length (p42) and truncated (p30) C/EBPα proteins with distinct functions can be formed. Start site B is out of frame and determines whether A or B is accessible for translation, hence producing either p42 or p30 C/EBPα proteins. (b) Schematic representation showing three important signaling pathways for the translation control of CEBPA messenger RNA: (1) the pathway leading to activation of the eukaryotic initiation factors eIF2 and eIF2B, which is counteracted by hnRNPE2, (2) the pathway of mTOR and eukaryotic initiation factor 4E (eIF4E), which is inhibited by 4E-BP1, and (3) the pathway leading to calreticulin (CRT) expression, a protein that binds to a double-stranded RNA loop and prevents the translation of full length C/EBPα proteins. Abbreviations: 5′ TOP: 5′ tract of pyrimidine; mTOR: mammalian target of rapamycin.
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fig1: (a) Simplified scheme of the CEBPA mRNA. Due to alternative translation start sites (A, B, and C), full length (p42) and truncated (p30) C/EBPα proteins with distinct functions can be formed. Start site B is out of frame and determines whether A or B is accessible for translation, hence producing either p42 or p30 C/EBPα proteins. (b) Schematic representation showing three important signaling pathways for the translation control of CEBPA messenger RNA: (1) the pathway leading to activation of the eukaryotic initiation factors eIF2 and eIF2B, which is counteracted by hnRNPE2, (2) the pathway of mTOR and eukaryotic initiation factor 4E (eIF4E), which is inhibited by 4E-BP1, and (3) the pathway leading to calreticulin (CRT) expression, a protein that binds to a double-stranded RNA loop and prevents the translation of full length C/EBPα proteins. Abbreviations: 5′ TOP: 5′ tract of pyrimidine; mTOR: mammalian target of rapamycin.

Mentions: In general, two mechanisms can be involved in translation control: “global” and “selective”. Global control acts on all mRNAs in a nonspecific manner, whereas selective translation regulation targets a specific subset of mRNAs. These specific mRNAs often have cis-regulatory sequences that sense subtle changes in the activity of the translation machinery or form loops that affect the accessibility of the appropriate translation start sites. It is now well documented that the CEBPA mRNA can be expressed as a full length protein (p42) or a truncated form (p30) [6, 7, 14–18]. The p42/p30 ratio is predominantly controlled at the translational level [15–18]. Full lengths (p42) and truncated (p30) C/EBPα proteins are generated from one single 5′ tract of pyrimidine (5′ TOP) CEBPA mRNA (Figure 1(a)). Three important signaling pathways regulate the translation of 5′ TOP messengers (Figure 1(b)). The first is the ubiquitous eukaryotic initiation factor 2 (eIF2). The second is leading to activation of mammalian target of rapamycin (mTOR) and subsequent activation of eukaryotic initiation factor 4E (eIF4E). These pathways are stringently controlled by specific inhibitory proteins, including hnRNPE2, which interferes with translation initiation, and 4E-BP1, a protein that prevents ribosomal scanning [19]. The third level of control of CEBPA mRNA translation is found in a cis-regulatory double-stranded RNA loop, which provides a docking site for calreticulin. When calreticulin is bound to this sequence, translation of the full length C/EBPα (p42) is reduced [20].


Calreticulin is a negative regulator of bronchial smooth muscle cell proliferation.

Miglino N, Roth M, Lardinois D, Tamm M, Borger P - J Allergy (Cairo) (2012)

(a) Simplified scheme of the CEBPA mRNA. Due to alternative translation start sites (A, B, and C), full length (p42) and truncated (p30) C/EBPα proteins with distinct functions can be formed. Start site B is out of frame and determines whether A or B is accessible for translation, hence producing either p42 or p30 C/EBPα proteins. (b) Schematic representation showing three important signaling pathways for the translation control of CEBPA messenger RNA: (1) the pathway leading to activation of the eukaryotic initiation factors eIF2 and eIF2B, which is counteracted by hnRNPE2, (2) the pathway of mTOR and eukaryotic initiation factor 4E (eIF4E), which is inhibited by 4E-BP1, and (3) the pathway leading to calreticulin (CRT) expression, a protein that binds to a double-stranded RNA loop and prevents the translation of full length C/EBPα proteins. Abbreviations: 5′ TOP: 5′ tract of pyrimidine; mTOR: mammalian target of rapamycin.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig1: (a) Simplified scheme of the CEBPA mRNA. Due to alternative translation start sites (A, B, and C), full length (p42) and truncated (p30) C/EBPα proteins with distinct functions can be formed. Start site B is out of frame and determines whether A or B is accessible for translation, hence producing either p42 or p30 C/EBPα proteins. (b) Schematic representation showing three important signaling pathways for the translation control of CEBPA messenger RNA: (1) the pathway leading to activation of the eukaryotic initiation factors eIF2 and eIF2B, which is counteracted by hnRNPE2, (2) the pathway of mTOR and eukaryotic initiation factor 4E (eIF4E), which is inhibited by 4E-BP1, and (3) the pathway leading to calreticulin (CRT) expression, a protein that binds to a double-stranded RNA loop and prevents the translation of full length C/EBPα proteins. Abbreviations: 5′ TOP: 5′ tract of pyrimidine; mTOR: mammalian target of rapamycin.
Mentions: In general, two mechanisms can be involved in translation control: “global” and “selective”. Global control acts on all mRNAs in a nonspecific manner, whereas selective translation regulation targets a specific subset of mRNAs. These specific mRNAs often have cis-regulatory sequences that sense subtle changes in the activity of the translation machinery or form loops that affect the accessibility of the appropriate translation start sites. It is now well documented that the CEBPA mRNA can be expressed as a full length protein (p42) or a truncated form (p30) [6, 7, 14–18]. The p42/p30 ratio is predominantly controlled at the translational level [15–18]. Full lengths (p42) and truncated (p30) C/EBPα proteins are generated from one single 5′ tract of pyrimidine (5′ TOP) CEBPA mRNA (Figure 1(a)). Three important signaling pathways regulate the translation of 5′ TOP messengers (Figure 1(b)). The first is the ubiquitous eukaryotic initiation factor 2 (eIF2). The second is leading to activation of mammalian target of rapamycin (mTOR) and subsequent activation of eukaryotic initiation factor 4E (eIF4E). These pathways are stringently controlled by specific inhibitory proteins, including hnRNPE2, which interferes with translation initiation, and 4E-BP1, a protein that prevents ribosomal scanning [19]. The third level of control of CEBPA mRNA translation is found in a cis-regulatory double-stranded RNA loop, which provides a docking site for calreticulin. When calreticulin is bound to this sequence, translation of the full length C/EBPα (p42) is reduced [20].

Bottom Line: In the presence of 5% FBS, the p42/p30 ratio significantly decreased (n = 3, P < 0.05) and coincided with BSM cell proliferation.High levels of calreticulin were associated with a decreased p42/p30 isoform ratio.Modulation of calreticulin levels may provide a novel target to reduce BSM remodeling.

View Article: PubMed Central - PubMed

Affiliation: Departments of Biomedicine, Pulmonary Cell Research, and Thorax Surgery, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland.

ABSTRACT
Background. Calreticulin controls the C/EBPαp42/p30 at the translational level trough a cis-regulatory CNG rich loop in the CEBPA mRNA. We determined the effects of steroids and long-acting beta-agonists on the p42/p30 ratio and on calreticulin expression in primary human bronchial smooth muscle (BSM) cells. Methods. The effects of budesonide (10(-8) M) and formoterol (10(-8) M) were studied in BSM cells pre-treated with siRNA targeting calreticulin. The expression of C/EBPα and calreticulin was determined by immuno-blotting. Automated cell counts were performed to measure proliferation. Results. All tested BSM cell lines (n = 5) expressed C/EBPα and calreticulin. In the presence of 5% FBS, the p42/p30 ratio significantly decreased (n = 3, P < 0.05) and coincided with BSM cell proliferation. High levels of calreticulin were associated with a decreased p42/p30 isoform ratio. FBS induced the expression of calreticulin (n = 3, P < 0.05), which was further increased by formoterol. siRNA targeting calreticulin increased the p42/p30 ratio in non-stimulated BSM cells and significantly inhibited the proliferation of PDGF-BB-stimulated BSM cells (n = 5, P < 0.05). Neither budesonide nor formoterol restored the p42 isoform expression. Conclusions. Our data show calreticulin is a negative regulator of C/EBPα protein expression in BSM cells. Modulation of calreticulin levels may provide a novel target to reduce BSM remodeling.

No MeSH data available.