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OSP/claudin-11 forms a complex with a novel member of the tetraspanin super family and beta1 integrin and regulates proliferation and migration of oligodendrocytes.

Tiwari-Woodruff SK, Buznikov AG, Vu TQ, Micevych PE, Chen K, Kornblum HI, Bronstein JM - J. Cell Biol. (2001)

Bottom Line: Overexpression of OSP/claudin-11 or OAP-1 induced proliferation in an oligodendrocyte cell line.Anti-OAP-1, anti-OSP/claudin-11, and anti-beta1 integrin antibodies inhibited migration of primary oligodendrocytes, and migration was impaired in OSP/claudin-11-deficient primary oligodendrocytes.These data suggest a role for OSP/claudin-11, OAP-1, and beta1 integrin complex in regulating proliferation and migration of oligodendrocytes, a process essential for normal myelination and repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of California at Los Angeles School of Medicine, Los Angeles, California 90095, USA.

ABSTRACT
Oligodendrocyte-specific protein (OSP)/claudin-11 is a major component of central nervous system myelin and forms tight junctions (TJs) within myelin sheaths. TJs are essential for forming a paracellular barrier and have been implicated in the regulation of growth and differentiation via signal transduction pathways. We have identified an OSP/claudin-11-associated protein (OAP)1, using a yeast two-hybrid screen. OAP-1 is a novel member of the tetraspanin superfamily, and it is widely expressed in several cell types, including oligodendrocytes. OAP-1, OSP/claudin-11, and beta1 integrin form a complex as indicated by coimmunoprecipitation and confocal immunocytochemistry. Overexpression of OSP/claudin-11 or OAP-1 induced proliferation in an oligodendrocyte cell line. Anti-OAP-1, anti-OSP/claudin-11, and anti-beta1 integrin antibodies inhibited migration of primary oligodendrocytes, and migration was impaired in OSP/claudin-11-deficient primary oligodendrocytes. These data suggest a role for OSP/claudin-11, OAP-1, and beta1 integrin complex in regulating proliferation and migration of oligodendrocytes, a process essential for normal myelination and repair.

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Overexpression of OSP/claudin-11 and OAP-1 increases proliferation in CIMO cells. (A) Northern blot of CIMO cells grown at 37°C (lanes 1 and 3) and 33°C (lanes 2 and 4) probed with a 32P-labeled 2-kb cDNA of OSP/claudin-11 (lanes 1 and 2) and 1.9-kB cDNA of OAP-1 (lanes 3 and 4). Both OSP/claudin-11 and OAP-1 expression is increased at 33°C during proliferation. (B) Western blot of CIMO cells transfected with control vector (lanes 1 and 3) and OSP (lanes 2 and 4) probed with anti-OSP antibody at 33°C (lanes 1 and 2) and 37°C (lanes 3 and 4). (Bottom panel) Western blot of CIMO cells transfected with OAP-1 (lane 1) and control vector (lane 2) at 37°C probed with anti–OAP-1 antibody confirms increased expression in these stable transfectents. (C) [3H]Thymidine incorporation into DNA of CIMO cells transfected with control vectors (pBabe and pcDNA) and OSP/claudin-11, antisense OSP/claudin-11, PLP, MBP, and OAP-1 constructs were observed under nonpermissive (37°C, minus Iγ) and permissive conditions (33°C, plus Iγ). OSP/claudin-11–transfected cells had a significant effect on proliferation at 33°C, whereas OAP-1 had a more generalized effect at both 33°C and 37°C. Control CIMO cells incorporated 4,000 ± 800 cpm/well/12 h, similar to the control vector transfected CIMO cells (average ± SEM, n = 24–32; 500 cells/well, each well counted in triplicates). OSP/claudin-11 versus pBabe vector at 37°C (***P < 0.0005), and OAP-1 versus pcDNA vector at 33°C and 37°C (**P < 0.005) (Student's t test).
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Figure 7: Overexpression of OSP/claudin-11 and OAP-1 increases proliferation in CIMO cells. (A) Northern blot of CIMO cells grown at 37°C (lanes 1 and 3) and 33°C (lanes 2 and 4) probed with a 32P-labeled 2-kb cDNA of OSP/claudin-11 (lanes 1 and 2) and 1.9-kB cDNA of OAP-1 (lanes 3 and 4). Both OSP/claudin-11 and OAP-1 expression is increased at 33°C during proliferation. (B) Western blot of CIMO cells transfected with control vector (lanes 1 and 3) and OSP (lanes 2 and 4) probed with anti-OSP antibody at 33°C (lanes 1 and 2) and 37°C (lanes 3 and 4). (Bottom panel) Western blot of CIMO cells transfected with OAP-1 (lane 1) and control vector (lane 2) at 37°C probed with anti–OAP-1 antibody confirms increased expression in these stable transfectents. (C) [3H]Thymidine incorporation into DNA of CIMO cells transfected with control vectors (pBabe and pcDNA) and OSP/claudin-11, antisense OSP/claudin-11, PLP, MBP, and OAP-1 constructs were observed under nonpermissive (37°C, minus Iγ) and permissive conditions (33°C, plus Iγ). OSP/claudin-11–transfected cells had a significant effect on proliferation at 33°C, whereas OAP-1 had a more generalized effect at both 33°C and 37°C. Control CIMO cells incorporated 4,000 ± 800 cpm/well/12 h, similar to the control vector transfected CIMO cells (average ± SEM, n = 24–32; 500 cells/well, each well counted in triplicates). OSP/claudin-11 versus pBabe vector at 37°C (***P < 0.0005), and OAP-1 versus pcDNA vector at 33°C and 37°C (**P < 0.005) (Student's t test).

Mentions: OAP-1 is widely expressed in brain and other tissues. Northern blot analysis revealed a 1.9-kb mRNA present in all the tissues tested, including spinal cord, brain, testes, skeletal muscle, heart, kidney, spleen, and lung (Fig. 2 A). A second band of ∼4 kb was also present in much lower levels. This may represent an alternate transcript or another gene product with high homology to OAP-1. Northern blot analysis on RNA from purified primary mouse oligodendrocyte cultures and from the CIMO cell line confirmed the presence of OAP-1 expression in oligodendrocytes (Fig. 2 B and see Fig. 7 A). Western blot analysis revealed a single band of ∼31 kD in O2A oligodendrocyte progenitor cells (Fig. 2 C). Two bands of ∼31 and 60 kD were present in mature oligodendrocytes and mouse brain homogenates, although under strong reducing conditions (20% β-mercaptoethanol and 50 mM DTT) only the 31-kD band was apparent. The band was eliminated by preincubation of the OAP-1 antibody with peptide (Fig. 2 C). These data suggest that the antibody specifically recognized OAP-1 protein and that the 60-kD band was a dimer of OAP-1 protein.


OSP/claudin-11 forms a complex with a novel member of the tetraspanin super family and beta1 integrin and regulates proliferation and migration of oligodendrocytes.

Tiwari-Woodruff SK, Buznikov AG, Vu TQ, Micevych PE, Chen K, Kornblum HI, Bronstein JM - J. Cell Biol. (2001)

Overexpression of OSP/claudin-11 and OAP-1 increases proliferation in CIMO cells. (A) Northern blot of CIMO cells grown at 37°C (lanes 1 and 3) and 33°C (lanes 2 and 4) probed with a 32P-labeled 2-kb cDNA of OSP/claudin-11 (lanes 1 and 2) and 1.9-kB cDNA of OAP-1 (lanes 3 and 4). Both OSP/claudin-11 and OAP-1 expression is increased at 33°C during proliferation. (B) Western blot of CIMO cells transfected with control vector (lanes 1 and 3) and OSP (lanes 2 and 4) probed with anti-OSP antibody at 33°C (lanes 1 and 2) and 37°C (lanes 3 and 4). (Bottom panel) Western blot of CIMO cells transfected with OAP-1 (lane 1) and control vector (lane 2) at 37°C probed with anti–OAP-1 antibody confirms increased expression in these stable transfectents. (C) [3H]Thymidine incorporation into DNA of CIMO cells transfected with control vectors (pBabe and pcDNA) and OSP/claudin-11, antisense OSP/claudin-11, PLP, MBP, and OAP-1 constructs were observed under nonpermissive (37°C, minus Iγ) and permissive conditions (33°C, plus Iγ). OSP/claudin-11–transfected cells had a significant effect on proliferation at 33°C, whereas OAP-1 had a more generalized effect at both 33°C and 37°C. Control CIMO cells incorporated 4,000 ± 800 cpm/well/12 h, similar to the control vector transfected CIMO cells (average ± SEM, n = 24–32; 500 cells/well, each well counted in triplicates). OSP/claudin-11 versus pBabe vector at 37°C (***P < 0.0005), and OAP-1 versus pcDNA vector at 33°C and 37°C (**P < 0.005) (Student's t test).
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Figure 7: Overexpression of OSP/claudin-11 and OAP-1 increases proliferation in CIMO cells. (A) Northern blot of CIMO cells grown at 37°C (lanes 1 and 3) and 33°C (lanes 2 and 4) probed with a 32P-labeled 2-kb cDNA of OSP/claudin-11 (lanes 1 and 2) and 1.9-kB cDNA of OAP-1 (lanes 3 and 4). Both OSP/claudin-11 and OAP-1 expression is increased at 33°C during proliferation. (B) Western blot of CIMO cells transfected with control vector (lanes 1 and 3) and OSP (lanes 2 and 4) probed with anti-OSP antibody at 33°C (lanes 1 and 2) and 37°C (lanes 3 and 4). (Bottom panel) Western blot of CIMO cells transfected with OAP-1 (lane 1) and control vector (lane 2) at 37°C probed with anti–OAP-1 antibody confirms increased expression in these stable transfectents. (C) [3H]Thymidine incorporation into DNA of CIMO cells transfected with control vectors (pBabe and pcDNA) and OSP/claudin-11, antisense OSP/claudin-11, PLP, MBP, and OAP-1 constructs were observed under nonpermissive (37°C, minus Iγ) and permissive conditions (33°C, plus Iγ). OSP/claudin-11–transfected cells had a significant effect on proliferation at 33°C, whereas OAP-1 had a more generalized effect at both 33°C and 37°C. Control CIMO cells incorporated 4,000 ± 800 cpm/well/12 h, similar to the control vector transfected CIMO cells (average ± SEM, n = 24–32; 500 cells/well, each well counted in triplicates). OSP/claudin-11 versus pBabe vector at 37°C (***P < 0.0005), and OAP-1 versus pcDNA vector at 33°C and 37°C (**P < 0.005) (Student's t test).
Mentions: OAP-1 is widely expressed in brain and other tissues. Northern blot analysis revealed a 1.9-kb mRNA present in all the tissues tested, including spinal cord, brain, testes, skeletal muscle, heart, kidney, spleen, and lung (Fig. 2 A). A second band of ∼4 kb was also present in much lower levels. This may represent an alternate transcript or another gene product with high homology to OAP-1. Northern blot analysis on RNA from purified primary mouse oligodendrocyte cultures and from the CIMO cell line confirmed the presence of OAP-1 expression in oligodendrocytes (Fig. 2 B and see Fig. 7 A). Western blot analysis revealed a single band of ∼31 kD in O2A oligodendrocyte progenitor cells (Fig. 2 C). Two bands of ∼31 and 60 kD were present in mature oligodendrocytes and mouse brain homogenates, although under strong reducing conditions (20% β-mercaptoethanol and 50 mM DTT) only the 31-kD band was apparent. The band was eliminated by preincubation of the OAP-1 antibody with peptide (Fig. 2 C). These data suggest that the antibody specifically recognized OAP-1 protein and that the 60-kD band was a dimer of OAP-1 protein.

Bottom Line: Overexpression of OSP/claudin-11 or OAP-1 induced proliferation in an oligodendrocyte cell line.Anti-OAP-1, anti-OSP/claudin-11, and anti-beta1 integrin antibodies inhibited migration of primary oligodendrocytes, and migration was impaired in OSP/claudin-11-deficient primary oligodendrocytes.These data suggest a role for OSP/claudin-11, OAP-1, and beta1 integrin complex in regulating proliferation and migration of oligodendrocytes, a process essential for normal myelination and repair.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, University of California at Los Angeles School of Medicine, Los Angeles, California 90095, USA.

ABSTRACT
Oligodendrocyte-specific protein (OSP)/claudin-11 is a major component of central nervous system myelin and forms tight junctions (TJs) within myelin sheaths. TJs are essential for forming a paracellular barrier and have been implicated in the regulation of growth and differentiation via signal transduction pathways. We have identified an OSP/claudin-11-associated protein (OAP)1, using a yeast two-hybrid screen. OAP-1 is a novel member of the tetraspanin superfamily, and it is widely expressed in several cell types, including oligodendrocytes. OAP-1, OSP/claudin-11, and beta1 integrin form a complex as indicated by coimmunoprecipitation and confocal immunocytochemistry. Overexpression of OSP/claudin-11 or OAP-1 induced proliferation in an oligodendrocyte cell line. Anti-OAP-1, anti-OSP/claudin-11, and anti-beta1 integrin antibodies inhibited migration of primary oligodendrocytes, and migration was impaired in OSP/claudin-11-deficient primary oligodendrocytes. These data suggest a role for OSP/claudin-11, OAP-1, and beta1 integrin complex in regulating proliferation and migration of oligodendrocytes, a process essential for normal myelination and repair.

Show MeSH
Related in: MedlinePlus