Limits...
Cytokine-Like Factor 1, an Essential Facilitator of Cardiotrophin-Like Cytokine:Ciliary Neurotrophic Factor Receptor α Signaling and sorLA-Mediated Turnover.

Larsen JV, Kristensen AM, Pallesen LT, Bauer J, Vægter CB, Nielsen MS, Madsen P, Petersen CM - Mol. Cell. Biol. (2016)

Bottom Line: The site for CNTFRα enables CLF-1 to promote CLC:CNTFRα complex formation and signaling.The second site establishes a link between the endocytic receptor sorLA and the tripartite CLC:CLF-1:CNTFRα complex and allows sorLA to downregulate the CNTFRα pool in stimulated cells.Finally, sorLA may bind and concentrate the tripartite soluble CLC:CLF-1:CNTFRα complex on cell membranes and thus facilitate its signaling through gp130/LIFRβ.

View Article: PubMed Central - PubMed

Affiliation: The MIND Center, Department of Biomedicine, Aarhus University, Aarhus, Denmark jvl@biomed.au.dk cmp@biomed.au.dk.

No MeSH data available.


Related in: MedlinePlus

Uptake of CLC:CLF-1 is impaired in sorLA-deficient astrocytes and hippocampal neurons. (A) Astrocytes isolated from wt and from sorLA KO mice were incubated with His-tagged CLC:CLF-1 (40 nM at 37°C). The cells were then washed prior to fixation and finally permeabilized before staining with mouse anti-His and rabbit anti-sorLA antibodies. Alexa Fluor 488-conjugated donkey anti-mouse and Alexa Fluor 568-conjugated goat anti-rabbit antibodies were used as secondary antibodies. (B) Uptake of His-tagged CLC:CLF-1 in wt and sorLA KO mouse hippocampal neurons. The cells were incubated with CLC:CLF-1, fixed, and stained with anti-His and anti-sorLA antibodies as described above.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4836274&req=5

Figure 4: Uptake of CLC:CLF-1 is impaired in sorLA-deficient astrocytes and hippocampal neurons. (A) Astrocytes isolated from wt and from sorLA KO mice were incubated with His-tagged CLC:CLF-1 (40 nM at 37°C). The cells were then washed prior to fixation and finally permeabilized before staining with mouse anti-His and rabbit anti-sorLA antibodies. Alexa Fluor 488-conjugated donkey anti-mouse and Alexa Fluor 568-conjugated goat anti-rabbit antibodies were used as secondary antibodies. (B) Uptake of His-tagged CLC:CLF-1 in wt and sorLA KO mouse hippocampal neurons. The cells were incubated with CLC:CLF-1, fixed, and stained with anti-His and anti-sorLA antibodies as described above.

Mentions: Uptake of CLC:CLF-1 was confirmed in astrocytes, which exhibit endogenous expression of sorLA but little or no CNTFRα as determined by Western blotting and lack of response to CLC:CLF-1 stimulation (data not shown). Cultured astrocytes isolated from wt and sorLA-deficient (KO) mice were incubated (25 min) in the absence or presence of 40 nM His-tagged CLC:CLF-1 prior to staining with anti-His and anti-sorLA antibodies. As demonstrated in Fig. 4A, sorLA-deficient cells showed little or no uptake of CLC:CLF-1, whereas wt astrocytes presented a significant vesicular uptake, which to a large extent colocalized with sorLA. Thus, as determined by manual counting, wt astrocytes contained 11.44 ± 4.02 (mean ± SD; n = 16) CLC:CLF-1-positive vesicles per cell in contrast to the 1.25 ± 1.29 (n = 16) positive vesicles per cell found in sorLA-deficient astrocytes. A similar accumulation of ligand colocalizing with sorLA was observed in isolated wt hippocampal (mouse) neurons expressing sorLA but not in sorLA-deficient hippocampal neurons (Fig. 4B).


Cytokine-Like Factor 1, an Essential Facilitator of Cardiotrophin-Like Cytokine:Ciliary Neurotrophic Factor Receptor α Signaling and sorLA-Mediated Turnover.

Larsen JV, Kristensen AM, Pallesen LT, Bauer J, Vægter CB, Nielsen MS, Madsen P, Petersen CM - Mol. Cell. Biol. (2016)

Uptake of CLC:CLF-1 is impaired in sorLA-deficient astrocytes and hippocampal neurons. (A) Astrocytes isolated from wt and from sorLA KO mice were incubated with His-tagged CLC:CLF-1 (40 nM at 37°C). The cells were then washed prior to fixation and finally permeabilized before staining with mouse anti-His and rabbit anti-sorLA antibodies. Alexa Fluor 488-conjugated donkey anti-mouse and Alexa Fluor 568-conjugated goat anti-rabbit antibodies were used as secondary antibodies. (B) Uptake of His-tagged CLC:CLF-1 in wt and sorLA KO mouse hippocampal neurons. The cells were incubated with CLC:CLF-1, fixed, and stained with anti-His and anti-sorLA antibodies as described above.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4836274&req=5

Figure 4: Uptake of CLC:CLF-1 is impaired in sorLA-deficient astrocytes and hippocampal neurons. (A) Astrocytes isolated from wt and from sorLA KO mice were incubated with His-tagged CLC:CLF-1 (40 nM at 37°C). The cells were then washed prior to fixation and finally permeabilized before staining with mouse anti-His and rabbit anti-sorLA antibodies. Alexa Fluor 488-conjugated donkey anti-mouse and Alexa Fluor 568-conjugated goat anti-rabbit antibodies were used as secondary antibodies. (B) Uptake of His-tagged CLC:CLF-1 in wt and sorLA KO mouse hippocampal neurons. The cells were incubated with CLC:CLF-1, fixed, and stained with anti-His and anti-sorLA antibodies as described above.
Mentions: Uptake of CLC:CLF-1 was confirmed in astrocytes, which exhibit endogenous expression of sorLA but little or no CNTFRα as determined by Western blotting and lack of response to CLC:CLF-1 stimulation (data not shown). Cultured astrocytes isolated from wt and sorLA-deficient (KO) mice were incubated (25 min) in the absence or presence of 40 nM His-tagged CLC:CLF-1 prior to staining with anti-His and anti-sorLA antibodies. As demonstrated in Fig. 4A, sorLA-deficient cells showed little or no uptake of CLC:CLF-1, whereas wt astrocytes presented a significant vesicular uptake, which to a large extent colocalized with sorLA. Thus, as determined by manual counting, wt astrocytes contained 11.44 ± 4.02 (mean ± SD; n = 16) CLC:CLF-1-positive vesicles per cell in contrast to the 1.25 ± 1.29 (n = 16) positive vesicles per cell found in sorLA-deficient astrocytes. A similar accumulation of ligand colocalizing with sorLA was observed in isolated wt hippocampal (mouse) neurons expressing sorLA but not in sorLA-deficient hippocampal neurons (Fig. 4B).

Bottom Line: The site for CNTFRα enables CLF-1 to promote CLC:CNTFRα complex formation and signaling.The second site establishes a link between the endocytic receptor sorLA and the tripartite CLC:CLF-1:CNTFRα complex and allows sorLA to downregulate the CNTFRα pool in stimulated cells.Finally, sorLA may bind and concentrate the tripartite soluble CLC:CLF-1:CNTFRα complex on cell membranes and thus facilitate its signaling through gp130/LIFRβ.

View Article: PubMed Central - PubMed

Affiliation: The MIND Center, Department of Biomedicine, Aarhus University, Aarhus, Denmark jvl@biomed.au.dk cmp@biomed.au.dk.

No MeSH data available.


Related in: MedlinePlus