Fibronectin-keratinocyte interactions during terminal differentiation

36

Ceil Biology International

HUMANHOX GENES ARE SEQUENTIALLY S125 ACTIVATED BY RETINOIC ACID Edoardo Boncinelli, Antonio Simeone, Dario Acampora. International Institute of Genetics & Biophysics, CNR, Naples, Italy. Istituto H.S. Raffaele. Fulvio Mavilio. Milan, Italy. acid Retinoic specifically (-1 induces human HOX gene expression in the carcinoma emhryonal cell line NTZ/Dl. We now show that the 9 genes of the HOX2 locus are differentially activated ' NT2/Dl cells exposed to RA concentratioki ranging from 1OnM to low. Genes located in the 3' half of the locus are induced at peak levels by 1OnM RA, whereas a concentration of lfl to low is required to fully activate 5' genes. Upon exposure to either high or low RA concentrations, HOX2 genes are sequentially activated, within hours or days, in the 3’ to 5’ direction along the locus. Activation of early responding genes does not require protein synthesis and is not primarily hue to stabilization of preexisting mRNA. These data show that RA activates-the expression of human HOX2 genes in vitro in a sequential order which is colinear with both their 3' to 5' order in the cluster and the spatially-restricted expression pattern along the embryonic anteroposterior body axis. The same activation pattern was observed for responding genes of the four HOX loci. Conversely, the four most 5' genes of HOX4 are expressed in untreated NTZ/Dl cells and sequentially downregulated after RA addition.

s127

MICROENVIRONMENT CONTROL OF IN UTRO CHONDROGENESIS

R. Cancedda, P. Castagnola, F. Descalzi, B. Dozin, P. Manduca, R. Quarto, L. Abelmoschi, G. Campanile and F. Rossi. lstituto Nazionale per la Ricerca sul Cancro, Genova, bly. When transferred into suspension culture on agarose-coated dishes, chick embryo prechondrogenic cell assumea chondrocyte phenotype and continue their

maturationtothehypertrophicstage.Duringthisprocess thesynthesisofcollagentype Iisrepressedwhilethegenes encoding collagens 11, IX and X are progressively activated. Additional markers expressed by in vitro differentiating chondrocytes at different stages of development has been evidencedin our laboratory. These

markers include a low molecular weight protein, named Ch21.Thecomplete aminoacidsequenceofthe Ch21has beendeterminedandtheproteinhasbeenassignedtothe superfamily of low molecular weight proteins sharing a basic framework for the binding and transport of small hydrophobic molecules. Addition of ascorbic acid to the

suspension culture promotes cell aggregation and extracellular matrix organization, giving rise to a structure resemblig in viva endochondral cartilage. At later Stage of culture, the conversion of hypertrophic chondrocytes to cells expressing bone markers has also been observed. The constitutive expression of the myc oncogene inhibits

theterminaldifferentiationofchondrocytesi.e.activation oftype Xcollagenexpression.

Reports, Vol. 14, Abstracts Supplement

1990

Fiona M. Watt, Josephine C. Adamsand Linda J. Nicholson. Keratinocyte Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WCZA 3PX, England. In humanepidermis, proliferation of keratinocytes is largely confined to the basal layer, which is in contact with the basement

membrane,and cells undergo terminal differentiation

as they move upwards through the

suprabasal layers.

Humankeratinocytes can be

grown in culture under conditions in which the cells form stratified colonies with the same basic

organisation as the epidermis. Such cultures provide a useful experimental model for investigating interactions assembly.

the role of cell-extracellular in differentiation and tissue

During terminal differentiation

keratinocytes

lose adhesiveness

matrix

in culture to fibronectin,

laminin and collagen types I and IV. The adhesive changes precede, by several hours,loss of the 13261 aat

and a5B1 integrins

from the cell

surface.

Keratinocyte adhesion to fibronectin is mediated by the a5B1 integrin and the decrease in adhesion of intact cells to fibronectin is correlated with a

decrease in affinity of ~~61 receptors for fibronectin. Modulation of integrin function

early in terminal differentiation may therefore be the primary event in determining cell migration out of the basal layer and occurs, in the case of asBl,at two levels: a decrease in ligand binding affinity, followed by receptor loss from the cell surface which is correlated with a decrease in abundance of the subunit mRNAs.

S128 THE RCN&&UN&;;4

MYOBLAST

Klaus von der Mark, He1 a von der Mark and K Simon Goodman. Max-Panck-Society, Clinical ResearchUnits for Rheumatology at the University of Erlangen-N&&erg, FRG Lamhin has been shown to affect various aspects of myogenic differentiation of mouse myoblasts: It promotes proliferation, motility, elongation and expression of muscle specific proteins and finally fusion. For adhesion and migration on lam&n m oblasts primarily utilize. the Elastase fragment El which corresponds to the lo armoflamini& wgle some adhesion occurs on % e El-x fragment (the center of the cross),but no migration. In order to identify the myoblast receptors involved in laminin recognition, detergent extracts of myoblasts were passed over a EH!5-latninin-Sepharose according to Gehlsen et al. (1988), and the inte ‘ns were eluted w$h EPTA. Analysis by ,SpS-Pr GE :z: ci I%‘%~ $?%+y%%%!d : ti$ a-subunit isolated from MeWo-melanoma cells b Kramer and Marks (1989). Neither a3- nor a Bsubunits bound, to the EHS-laminin column, a6/fi 1 1smvolved in B16 ccjl adhesion to almu E?Pfragment. Analysis of VIA mtegrms from cell surface-iodinated myoblasts by immunopreci itation confhmed the absence of a6 from myobSasts, su~tiugthatthenovelaR/~l-complex is responsible r myoblast-E8 interaction.