Identification of MANF as a protein interacting wi
Post# of 30028
Posted On: 12/30/2014 11:38:22 PM
Identification of MANF as a protein interacting with RTN1-C.
Abstract
Reticulons (RTNs) constitute a family of endoplasmic reticulum (ER)-associated proteins with a reticular distribution. Recently, evidence has shown that they exert a cancer-specific proapoptotic function via interaction or modulation of specific proteins. Such evidence is particularly pertinent for the RTN1-C family member. In order to identify proteins that interact with RTN1-C, the yeast two-hybrid system and regular molecular biological techniques were used to screen the human fetal brain cDNA library. As a result, seven RTN1-C interacting proteins including Homo sapiens mesencephalic astrocyte-derived neurotrophic factor (MANF) were obtained. The interactions between RTN1-C and its interacting proteins were confirmed by using both β-galactosidase assay and growth test in selective media. Moreover, the MANF/RTN1-C interaction was confirmed in vitro by glutathione S-transferase pull-down and in vivo by immunoprecipitation assays. By immunofluorescence assay, it was found that MANF co-localized with RTN1-C in the ER. Knockdown of RTN1-C reduced the localization of MANF in the ER. These results provide clues to further explore the function of RTN1-C and MANF in neurodegenerative diseases and cancer.
© The Author 2014. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
Abstract
Reticulons (RTNs) constitute a family of endoplasmic reticulum (ER)-associated proteins with a reticular distribution. Recently, evidence has shown that they exert a cancer-specific proapoptotic function via interaction or modulation of specific proteins. Such evidence is particularly pertinent for the RTN1-C family member. In order to identify proteins that interact with RTN1-C, the yeast two-hybrid system and regular molecular biological techniques were used to screen the human fetal brain cDNA library. As a result, seven RTN1-C interacting proteins including Homo sapiens mesencephalic astrocyte-derived neurotrophic factor (MANF) were obtained. The interactions between RTN1-C and its interacting proteins were confirmed by using both β-galactosidase assay and growth test in selective media. Moreover, the MANF/RTN1-C interaction was confirmed in vitro by glutathione S-transferase pull-down and in vivo by immunoprecipitation assays. By immunofluorescence assay, it was found that MANF co-localized with RTN1-C in the ER. Knockdown of RTN1-C reduced the localization of MANF in the ER. These results provide clues to further explore the function of RTN1-C and MANF in neurodegenerative diseases and cancer.
© The Author 2014. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
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