CD44 modulates Smad1 activation in the BMP-7 signaling pathway.

Bone morphogenetic protein 7 (BMP-7) regulates cellular metabolism in embryonic and adult tissues. Signal transduction occurs through the activation of intracellular Smad proteins. In this paper, using a yeast two-hybrid screen, Smad1 was found to interact with the cytoplasmic domain of CD44, a receptor for the extracellular matrix macromolecule hyaluronan. Coimmunoprecipitation experiments confirmed the interaction of Smad1 with full-length CD44-interactions that did not occur when CD44 receptors truncated within the cytoplasmic domain were tested. Chondrocytes overexpressing a truncated CD44 on a background of endogenous full-length CD44 no longer exhibited Smad1 nuclear translocation upon BMP-7 stimulation. Further, pretreatment of chondrocytes with Streptomyces hyaluronidase to disrupt extracellular hyaluronan-cell interactions inhibited BMP-7-mediated Smad1 phosphorylation, nuclear translocation of Smad1 or Smad4, and SBE4-luciferase reporter activation. These results support a functional link between the BMP signaling cascade and CD44. Thus, changes in hyaluronan-cell interactions may serve as a means to modulate cellular responsiveness to BMP.

Radial glia express aromatase in the injured zebra finch brain.

Estrogens have neurotrophic and neuroprotective properties. The synthesis of estrogen occurs via the expression of aromatase. Previous studies have shown that injury to the vertebrate brain results in a rapid and dramatic up-regulation of aromatase expression in astrocytes around the lesion. As part of experiments examining injury-induced glial aromatization, we identified aromatase in radial glia of the zebra finch brain. Adult female zebra finches received a penetrating injury to the right hippocampus. Twenty-four hours after lesioning, birds were administered bromodeoxyuridine (BrdU) and sacrificed 2 hours, 1 day, or 7 days later. We determined the distribution of aromatase and BrdU labeling by using immunocytochemistry. Radial aromatase was localized to cells lining the lateral ventricle adjacent to the lesioned hippocampus. Injury also induced a dramatic accumulation of newly generated cells labeled with BrdU around the lesion. BrdU labeling was strongly associated with aromatase-positive radial fibers, suggesting the migration of newly generated cells along these fibers. In the songbird brain, estrogen supports neuronal recruitment and promotes the survival and addition of new neurons. The presence of aromatase in radial glia provides a mechanism of estrogen delivery to postmitotic cells. Radial aromatization may be a key feature in the repair of the vertebrate brain following neural injury.

A requirement for the CD44 cytoplasmic domain for hyaluronan binding, pericellular matrix assembly, and receptor-mediated endocytosis in COS-7 cells.

CD44-negative COS-7 cells were transfected with expression constructs for CD44H (the predominant CD44 isoform), CD44E (epithelial isoform), or truncation mutant derivatives lacking the carboxyl-terminal 67 amino acids of the cytoplasmic domain, CD44HDelta67 and CD44EDelta67. The truncation mutant CD44HDelta67 is identical to a naturally occurring alternatively spliced "short tail" CD44 isoform (CD44st), which incorporates exon 19 in place of exon 20. CD44st lacks intracellular signaling motifs as well as protein domains necessary for interaction with cytoskeletal components. Transfection of COS-7 cells with each construct yielded equivalent levels of mRNA expression, whereas no CD44 expression was observed in parental, nontransfected COS-7 cells. Western analysis and immunostaining of COS-7 transfectants confirmed CD44 protein expression of the truncation mutant derivatives. COS-7 cells transfected with CD44H or CD44E gained the capacity to bind fluorescein-conjugated HA (fl-HA) and assemble HA-dependent pericellular matrices in the presence of exogenously added HA and proteoglycan. In addition, the CD44H- and CD44E-transfected cells were able to internalize surface-bound fl-HA. COS-7 cells transfected with the vector alone or with either of the mutant CD44 isoforms, CD44HDelta67 or CD44EDelta67, did not exhibit the capacity to assemble pericellular matrices or to bind and internalize the fl-HA. Cotransfection of CD44Delta67 mutants together with CD44H reduced the size of the HA-dependent pericellular matrices. Transfection of bovine articular chondrocytes with CD44Delta67 also inhibited pericellular matrix assembly. Collectively, these results indicate an obligatory requirement for the CD44 receptor cytoplasmic domain for ligand (HA) binding, formation and retention of the pericellular matrix, as well as CD44-mediated endocytosis of HA. In addition, the results suggest a potential regulatory role for the differentially expressed alternatively spliced short tail CD44 isoform.