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CXCR7 protein expression in human adult brain and differentiated neurons.

Shimizu S, Brown M, Sengupta R, Penfold ME, Meucci O - PLoS ONE (2011)

Bottom Line: Furthermore, confocal microscopy and binding assays in cultured neurons show that CXCR7 protein is mainly located into cytoplasm, while little to no protein expression is found on neuronal plasma membrane.Interestingly, specific CXCR7 ligands that inhibit CXCL12 binding to CXCR7 do not alter CXCR4-activated survival signaling (pERK/pAkt) in rat cortical neurons.Overall, these results implicate CXCR4 as the main CXCL12 signaling receptor on the surface of differentiated neurons and suggest that CXCR7 may interact with CXCR4 at the intracellular level, possibly affecting CXCR4 trafficking and/or coupling to other proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Physiology, Drexel University College of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT

Background: CXCR7 and CXCR4 are receptors for the chemokine CXCL12, which is involved in essential functions of the immune and nervous systems. Although CXCR7 transcripts are widely expressed throughout the central nervous system, little is known about its protein distribution and function in the adult brain. To evaluate its potential involvement in CXCL12/CXCR4 signaling in differentiated neurons, we studied CXCR7 protein expression in human brain and cultured neurons.

Methodology/principal findings: Immunohistochemistry and RT-PCR analyses of cortex and hippocampus from control and HIV-positive subjects provided the first evidence of CXCR7 protein expression in human adult neurons, under normal and pathological conditions. Furthermore, confocal microscopy and binding assays in cultured neurons show that CXCR7 protein is mainly located into cytoplasm, while little to no protein expression is found on neuronal plasma membrane. Interestingly, specific CXCR7 ligands that inhibit CXCL12 binding to CXCR7 do not alter CXCR4-activated survival signaling (pERK/pAkt) in rat cortical neurons. Neuronal CXCR7 co-localizes to some extent with the endoplasmic reticulum marker ERp29, but not with early/late endosome markers. Additionally, large areas of overlap are detected in the intracellular pattern of CXCR7 and CXCR4 expression.

Conclusions/significance: Overall, these results implicate CXCR4 as the main CXCL12 signaling receptor on the surface of differentiated neurons and suggest that CXCR7 may interact with CXCR4 at the intracellular level, possibly affecting CXCR4 trafficking and/or coupling to other proteins.

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Related in: MedlinePlus

CXCR7 protein expression in the human brain.Staining with 11G8 or ab72100 was performed on different human tissues,                            using either a tissue array containing samples from multiple normal                            organs or additional brain sections as described in the methods. Images                            from the multi-organ array are shown in the top panels (A to C; scale                            bars: 200 µm). Neuronal CXCR7 expression was observed in the gray                            matter (A) while no staining is observed in normal breast tissue (B), as                            expected. CXCR7 was also expressed in the kidney (C). Additionally,                            serial sections from human frontal cortex were stained with: (D) rabbit                            polyclonal antibody ab72100, (E) 11G8, or (F) H&E (details in                            methods). Similar neuronal staining was observed with both CXCR7                            antibodies. No staining was detected when the primary antibody was                            omitted (G) or replaced by the isotype control (H). Images in insets are                            higher magnification of D, E, and F. Scale bars: 400 µm                            (D–H), 50 µm for insets. NC = negative                            control.
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pone-0020680-g002: CXCR7 protein expression in the human brain.Staining with 11G8 or ab72100 was performed on different human tissues, using either a tissue array containing samples from multiple normal organs or additional brain sections as described in the methods. Images from the multi-organ array are shown in the top panels (A to C; scale bars: 200 µm). Neuronal CXCR7 expression was observed in the gray matter (A) while no staining is observed in normal breast tissue (B), as expected. CXCR7 was also expressed in the kidney (C). Additionally, serial sections from human frontal cortex were stained with: (D) rabbit polyclonal antibody ab72100, (E) 11G8, or (F) H&E (details in methods). Similar neuronal staining was observed with both CXCR7 antibodies. No staining was detected when the primary antibody was omitted (G) or replaced by the isotype control (H). Images in insets are higher magnification of D, E, and F. Scale bars: 400 µm (D–H), 50 µm for insets. NC = negative control.

Mentions: Next, a tissue microarray containing samples of various normal human organs, including the brain, was analyzed. Initial studies with 11G8 showed a typical neuronal staining in the cerebral gray matter (Figure 2A/E); CXCR7 staining was also seen in tubular formation in the kidney (Figure 2C), in line with a previous study [37], while normal breast tissue was negative as expected (Figure 2B). Staining in the gray matter was also confirmed by using another CXCR7 antibody (ab72100, Figure 2D) that recognizes distinct portions of the receptor, i.e. epitopes in the second extracellular loop as opposed to the N-terminus recognized by 11G8. Though both these antibodies indicated neuronal expression of CXCR7, we primarily used 11G8 to keep consistency with the rest of the data.


CXCR7 protein expression in human adult brain and differentiated neurons.

Shimizu S, Brown M, Sengupta R, Penfold ME, Meucci O - PLoS ONE (2011)

CXCR7 protein expression in the human brain.Staining with 11G8 or ab72100 was performed on different human tissues,                            using either a tissue array containing samples from multiple normal                            organs or additional brain sections as described in the methods. Images                            from the multi-organ array are shown in the top panels (A to C; scale                            bars: 200 µm). Neuronal CXCR7 expression was observed in the gray                            matter (A) while no staining is observed in normal breast tissue (B), as                            expected. CXCR7 was also expressed in the kidney (C). Additionally,                            serial sections from human frontal cortex were stained with: (D) rabbit                            polyclonal antibody ab72100, (E) 11G8, or (F) H&E (details in                            methods). Similar neuronal staining was observed with both CXCR7                            antibodies. No staining was detected when the primary antibody was                            omitted (G) or replaced by the isotype control (H). Images in insets are                            higher magnification of D, E, and F. Scale bars: 400 µm                            (D–H), 50 µm for insets. NC = negative                            control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3105114&req=5

pone-0020680-g002: CXCR7 protein expression in the human brain.Staining with 11G8 or ab72100 was performed on different human tissues, using either a tissue array containing samples from multiple normal organs or additional brain sections as described in the methods. Images from the multi-organ array are shown in the top panels (A to C; scale bars: 200 µm). Neuronal CXCR7 expression was observed in the gray matter (A) while no staining is observed in normal breast tissue (B), as expected. CXCR7 was also expressed in the kidney (C). Additionally, serial sections from human frontal cortex were stained with: (D) rabbit polyclonal antibody ab72100, (E) 11G8, or (F) H&E (details in methods). Similar neuronal staining was observed with both CXCR7 antibodies. No staining was detected when the primary antibody was omitted (G) or replaced by the isotype control (H). Images in insets are higher magnification of D, E, and F. Scale bars: 400 µm (D–H), 50 µm for insets. NC = negative control.
Mentions: Next, a tissue microarray containing samples of various normal human organs, including the brain, was analyzed. Initial studies with 11G8 showed a typical neuronal staining in the cerebral gray matter (Figure 2A/E); CXCR7 staining was also seen in tubular formation in the kidney (Figure 2C), in line with a previous study [37], while normal breast tissue was negative as expected (Figure 2B). Staining in the gray matter was also confirmed by using another CXCR7 antibody (ab72100, Figure 2D) that recognizes distinct portions of the receptor, i.e. epitopes in the second extracellular loop as opposed to the N-terminus recognized by 11G8. Though both these antibodies indicated neuronal expression of CXCR7, we primarily used 11G8 to keep consistency with the rest of the data.

Bottom Line: Furthermore, confocal microscopy and binding assays in cultured neurons show that CXCR7 protein is mainly located into cytoplasm, while little to no protein expression is found on neuronal plasma membrane.Interestingly, specific CXCR7 ligands that inhibit CXCL12 binding to CXCR7 do not alter CXCR4-activated survival signaling (pERK/pAkt) in rat cortical neurons.Overall, these results implicate CXCR4 as the main CXCL12 signaling receptor on the surface of differentiated neurons and suggest that CXCR7 may interact with CXCR4 at the intracellular level, possibly affecting CXCR4 trafficking and/or coupling to other proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Physiology, Drexel University College of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT

Background: CXCR7 and CXCR4 are receptors for the chemokine CXCL12, which is involved in essential functions of the immune and nervous systems. Although CXCR7 transcripts are widely expressed throughout the central nervous system, little is known about its protein distribution and function in the adult brain. To evaluate its potential involvement in CXCL12/CXCR4 signaling in differentiated neurons, we studied CXCR7 protein expression in human brain and cultured neurons.

Methodology/principal findings: Immunohistochemistry and RT-PCR analyses of cortex and hippocampus from control and HIV-positive subjects provided the first evidence of CXCR7 protein expression in human adult neurons, under normal and pathological conditions. Furthermore, confocal microscopy and binding assays in cultured neurons show that CXCR7 protein is mainly located into cytoplasm, while little to no protein expression is found on neuronal plasma membrane. Interestingly, specific CXCR7 ligands that inhibit CXCL12 binding to CXCR7 do not alter CXCR4-activated survival signaling (pERK/pAkt) in rat cortical neurons. Neuronal CXCR7 co-localizes to some extent with the endoplasmic reticulum marker ERp29, but not with early/late endosome markers. Additionally, large areas of overlap are detected in the intracellular pattern of CXCR7 and CXCR4 expression.

Conclusions/significance: Overall, these results implicate CXCR4 as the main CXCL12 signaling receptor on the surface of differentiated neurons and suggest that CXCR7 may interact with CXCR4 at the intracellular level, possibly affecting CXCR4 trafficking and/or coupling to other proteins.

Show MeSH
Related in: MedlinePlus