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SNX16 negatively regulates the migration and tumorigenesis of MCF-7 cells.

Zhang L, Qin D, Hao C, Shu X, Pei D - Cell Regen (Lond) (2013)

Bottom Line: Ectopic expression of SNX16 reduces the migration and the tumor formation activity of MCF-7 cells.Our results indicate that, in addition to the PI3P, there is a SNX23- and microtubule-dependent cargo transport pathway required for the proper subcellular distribution of SNX16.SNX16 plays a negative regulatory role during cell migration and tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Regenerative Biology, Chinese Academy of Sciences, and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Guangzhou, 510530 China.

ABSTRACT

Background: Sorting nexins are a large family of proteins that are associated with various components of the endosome system and they play many roles in processes such as endocytosis, intracellular protein trafficking and cell signaling. The subcellular distribution patterns of many of them remain controversial and their in vivo functions have not been characterized yet.

Results: We investigated the subcellular distribution and function of SNX16 in this study. SNX16 is detected on Rab5-positive endosomes localized adjacent to focal adhesions at cell cortex. Inhibition of SNX23, polymerization of microtubule filaments as well as the PI3-kinase all disrupt the cell cortex distribution of SNX16. Ectopic expression of SNX16 reduces the migration and the tumor formation activity of MCF-7 cells.

Conclusion: Our results indicate that, in addition to the PI3P, there is a SNX23- and microtubule-dependent cargo transport pathway required for the proper subcellular distribution of SNX16. SNX16 plays a negative regulatory role during cell migration and tumorigenesis.

No MeSH data available.


Related in: MedlinePlus

Subcellular distribution of SNX16. (A) The subcellular distribution of tagged SNX16 in MCF-7 cells. Cells were transfected with the indicated constructs and immunofluoresence staining was performed 48 hrs post transfection. Rab5 is an early endosome marker and it co-localizes with SNX16 at cell cortex (indicated by an arrow). The endogenous Paxillin is detected using a specific antibody and used to indicate the position of focal adhesions. (B) The cell cortex distribution of SNX16 is detectable in a variety of cell lines. (C) A home-made polyclonal antibody to human SNX16 can detect the ectopically expressed SNX16. (D) Immunofluoresence staining of endogenous SNX16 on frozen sections prepared from adult mouse heart. Pre-incubation of the sample with soluble SNX16 protein blocks the staining.
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Fig1: Subcellular distribution of SNX16. (A) The subcellular distribution of tagged SNX16 in MCF-7 cells. Cells were transfected with the indicated constructs and immunofluoresence staining was performed 48 hrs post transfection. Rab5 is an early endosome marker and it co-localizes with SNX16 at cell cortex (indicated by an arrow). The endogenous Paxillin is detected using a specific antibody and used to indicate the position of focal adhesions. (B) The cell cortex distribution of SNX16 is detectable in a variety of cell lines. (C) A home-made polyclonal antibody to human SNX16 can detect the ectopically expressed SNX16. (D) Immunofluoresence staining of endogenous SNX16 on frozen sections prepared from adult mouse heart. Pre-incubation of the sample with soluble SNX16 protein blocks the staining.

Mentions: SNX16 has been detected at various endosome compartments including early endosomes, late endosomes/lysosomes or recycling endosomes; however, the exact subcellular distribution of SNX16 appears to be cell line dependent [31–34]. We initially investigated the distribution of ectopic SNX16 (Flag- or GFP-tagged) in MCF-7 which is a commonly used cell line derived from human breast cancer. We found that, in addition to the perinuclear region of cytoplasm, SNX16 vesicles are accumulated at certain cell cortex (indicated by arrows in Figure 1A). These vesicles are Rab5-positive so they are likely to be early endosomes. This distinct distribution pattern of SNX16 prompted us to investigate whether or not it is related to the focal adhesions, where a cell is linked to the extracellular matrix. Paxillin is a focal adhesion-associated adaptor protein and it is used to indicate the position of focal adhesions. We found that the cell cortex fraction of SNX16 is always adjacent to the Paxillin staining signals but they usually do not co-localize with each other. So we conclude that SNX16 vesicles are accumulated near certain focal adhesions at the peripheral cytoplasm in MCF-7 cells.Figure 1


SNX16 negatively regulates the migration and tumorigenesis of MCF-7 cells.

Zhang L, Qin D, Hao C, Shu X, Pei D - Cell Regen (Lond) (2013)

Subcellular distribution of SNX16. (A) The subcellular distribution of tagged SNX16 in MCF-7 cells. Cells were transfected with the indicated constructs and immunofluoresence staining was performed 48 hrs post transfection. Rab5 is an early endosome marker and it co-localizes with SNX16 at cell cortex (indicated by an arrow). The endogenous Paxillin is detected using a specific antibody and used to indicate the position of focal adhesions. (B) The cell cortex distribution of SNX16 is detectable in a variety of cell lines. (C) A home-made polyclonal antibody to human SNX16 can detect the ectopically expressed SNX16. (D) Immunofluoresence staining of endogenous SNX16 on frozen sections prepared from adult mouse heart. Pre-incubation of the sample with soluble SNX16 protein blocks the staining.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4230745&req=5

Fig1: Subcellular distribution of SNX16. (A) The subcellular distribution of tagged SNX16 in MCF-7 cells. Cells were transfected with the indicated constructs and immunofluoresence staining was performed 48 hrs post transfection. Rab5 is an early endosome marker and it co-localizes with SNX16 at cell cortex (indicated by an arrow). The endogenous Paxillin is detected using a specific antibody and used to indicate the position of focal adhesions. (B) The cell cortex distribution of SNX16 is detectable in a variety of cell lines. (C) A home-made polyclonal antibody to human SNX16 can detect the ectopically expressed SNX16. (D) Immunofluoresence staining of endogenous SNX16 on frozen sections prepared from adult mouse heart. Pre-incubation of the sample with soluble SNX16 protein blocks the staining.
Mentions: SNX16 has been detected at various endosome compartments including early endosomes, late endosomes/lysosomes or recycling endosomes; however, the exact subcellular distribution of SNX16 appears to be cell line dependent [31–34]. We initially investigated the distribution of ectopic SNX16 (Flag- or GFP-tagged) in MCF-7 which is a commonly used cell line derived from human breast cancer. We found that, in addition to the perinuclear region of cytoplasm, SNX16 vesicles are accumulated at certain cell cortex (indicated by arrows in Figure 1A). These vesicles are Rab5-positive so they are likely to be early endosomes. This distinct distribution pattern of SNX16 prompted us to investigate whether or not it is related to the focal adhesions, where a cell is linked to the extracellular matrix. Paxillin is a focal adhesion-associated adaptor protein and it is used to indicate the position of focal adhesions. We found that the cell cortex fraction of SNX16 is always adjacent to the Paxillin staining signals but they usually do not co-localize with each other. So we conclude that SNX16 vesicles are accumulated near certain focal adhesions at the peripheral cytoplasm in MCF-7 cells.Figure 1

Bottom Line: Ectopic expression of SNX16 reduces the migration and the tumor formation activity of MCF-7 cells.Our results indicate that, in addition to the PI3P, there is a SNX23- and microtubule-dependent cargo transport pathway required for the proper subcellular distribution of SNX16.SNX16 plays a negative regulatory role during cell migration and tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Regenerative Biology, Chinese Academy of Sciences, and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Guangzhou, 510530 China.

ABSTRACT

Background: Sorting nexins are a large family of proteins that are associated with various components of the endosome system and they play many roles in processes such as endocytosis, intracellular protein trafficking and cell signaling. The subcellular distribution patterns of many of them remain controversial and their in vivo functions have not been characterized yet.

Results: We investigated the subcellular distribution and function of SNX16 in this study. SNX16 is detected on Rab5-positive endosomes localized adjacent to focal adhesions at cell cortex. Inhibition of SNX23, polymerization of microtubule filaments as well as the PI3-kinase all disrupt the cell cortex distribution of SNX16. Ectopic expression of SNX16 reduces the migration and the tumor formation activity of MCF-7 cells.

Conclusion: Our results indicate that, in addition to the PI3P, there is a SNX23- and microtubule-dependent cargo transport pathway required for the proper subcellular distribution of SNX16. SNX16 plays a negative regulatory role during cell migration and tumorigenesis.

No MeSH data available.


Related in: MedlinePlus