The alpha 1-alpha 6 subunits of integrins are characteristically expressed in distinct segments of developing and adult human nephron.

We studied the distribution of the alpha 1-alpha 6 subunits of beta 1 integrins in developing and adult human kidney using a panel of mAbs in indirect immunofluorescence microscopy. Uninduced mesenchyme displayed a diffuse immunoreactivity for only the alpha 1 integrin subunit. At the S-shaped body stage of nephron development, several of the alpha subunits were characteristically expressed in distinct fetal nephron segments, and the pattern was retained also in the adult nephron. Thus, the alpha 1 subunit was characteristically expressed in mesangial and endothelial cells, the alpha 2 in glomerular endothelium and distal tubules, the alpha 3 in podocytes, Bowman's capsule, and distal tubules, and the alpha 6 subunit basally in all tubules, and only transiently in podocytes during development. Unlike the alpha 3 and alpha 6 subunits, the alpha 2 subunit displayed an overall cell surface distribution in distal tubules. It was also distinctly expressed in glomerular endothelia during glomerulogenesis. The beta 4 subunit was expressed only in fetal collecting ducts, and hence the alpha 6 subunit seems to be complexed with the beta 1 rather than beta 4 subunit in human kidney. Of the two fibronectin receptor alpha subunits, alpha 4 and alpha 5, only the latter was expressed, confined to endothelia of developing and adult blood vessels, suggesting that these receptor complexes play a minor role during nephrogenesis. The present results suggest that distinct integrins play a role during differentiation of specific nephron segments. They also indicate that alpha 3 beta 1 and alpha 6 beta 1 integrin complexes may function as basement membrane receptors in podocytes and tubular epithelial cells.

A point mutation of integrin beta 1 subunit blocks binding of alpha 5 beta 1 to fibronectin and invasin but not recruitment to adhesion plaques.

A point mutation in a highly conserved region of the beta 1 subunit, Asp130 to Ala (D130A) substitution, abrogates the Arg-Gly-Asp (RGD)-dependent binding of alpha 5 beta 1 to fibronectin (FN) without disrupting gross structure or heterodimer assembly. The D130A mutation also interferes with binding to invasin, a ligand that lacks RGD sequence. In spite of the lack of detectable FN binding by alpha 5 beta 1(D130A), it was recruited to adhesion plaques formed on FN by endogenous hamster receptors. Thus, intact ligand binding function is not required for recruitment of alpha 5 beta 1 to adhesion plaques. Overexpression of beta 1(D130A) partially interfered with endogenous alpha 5 beta 1 function, thus defining a dominant negative beta 1 integrin mutation.

Distinct functions of integrin alpha and beta subunit cytoplasmic domains in cell spreading and formation of focal adhesions.

Integrin-mediated cell adhesion often results in cell spreading and the formation of focal adhesions. We exploited the capacity of recombinant human alpha IIb beta 3 integrin to endow heterologous cells with the ability to adhere and spread on fibrinogen to study the role of integrin cytoplasmic domains in initiation of cell spreading and focal adhesions. The same constructs were also used to analyze the role of the cytoplasmic domains in maintenance of the fidelity of the integrin repertoire at focal adhesions. Truncation mutants of the cytoplasmic domain of alpha IIb did not interfere with the ability of alpha IIb beta 3 to initiate cell spreading and form focal adhesions. Nevertheless, deletion of the alpha IIb cytoplasmic domain allowed indiscriminate recruitment of alpha IIb beta 3 to focal adhesions formed by other integrins. Truncation of the beta 3 subunit cytoplasmic domain abolished cell spreading mediated by alpha IIb beta 3 and also abrogated recruitment of alpha IIb beta 3 to focal adhesions. This truncation also dramatically impaired the ability of alpha IIb beta 3 to mediate the contraction of fibrin gels. In contrast, the beta 3 subunit cytoplasmic truncation did not reduce the fibrinogen binding affinity of alpha IIb beta 3. Thus, the integrin beta 3 subunit cytoplasmic domain is necessary and sufficient for initiation of cell spreading and focal adhesion formation. Further, the beta 3 cytoplasmic domain is required for the transmission of intracellular contractile forces to fibrin gels. The alpha subunit cytoplasmic domain maintains the fidelity of recruitment of the integrins to focal adhesions and thus regulates their repertoire of integrins.

Crystal structure of the alpha-actinin rod reveals an extensive torsional twist.

BACKGROUND: Alpha-actinin is a ubiquitously expressed protein found in numerous actin structures. It consists of an N-terminal actin binding domain, a central rod domain, and a C-terminal domain and functions as a homodimer to cross-link actin filaments. The rod domain determines the distance between cross-linked actin filaments and also serves as an interaction site for several cytoskeletal and signaling proteins. RESULTS: We report here the crystal structure of the alpha-actinin rod. The structure is a twisted antiparallel dimer that contains a conserved acidic surface. CONCLUSIONS: The novel features revealed by the structure allow prediction of the orientation of parallel and antiparallel cross-linked actin filaments in relation to alpha-actinin. The conserved acidic surface is a possible interaction site for several cytoplasmic tails of transmembrane proteins involved in the recruitment of alpha-actinin to the plasma membrane.

Expression of megakaryocytic and erythroid properties in human leukemic cells.

Previous studies have suggested that megakaryocytes and erythrocytes may share a common precursor cell. However, studies on the commitment to erythroid and megakaryocytic lineages have been hampered by the lack of suitable human leukemic cell lines having this kind of bipotential differentiation capability. We investigated the coexpression of megakaryocytic and erythroid markers in human leukemic cell lines as well as the capability of these cells to further differentiate upon exposure to differentiation-inducing agents. We report that the JK-1 cell line, previously characterized as a typically erythroid cell line with spontaneous differentiation to red cells, actually coexpressed megakaryocytic cell surface antigens and erythroid spectrins. We also report that the JK-1 cells could be induced to differentiate along the megakaryocytic lineage by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA). The other cell lines studied variably expressed megakaryocytic and erythroid antigens, the DAMI and CMK cells predominantly megakaryocytic properties and the T-33 and K562 cells some erythroid markers, whereas the HEL cells expressed markers for both lineages of differentiation. Our results suggest that the JK-1 cell line represents an immature cell population that has not yet been committed to either of the two lineages of differentiation. The JK-1 cell line might provide a useful tool for further studies on the transcriptional regulation of erythroid and megakaryocytic phenotypes and for studies on the commitment to these lineages of differentiation. Our results also suggest that the leukemic cell lines show a considerable plasticity in the expression of properties normally specific for distinct lineages of differentiation.

Tumor promoter and fibronectin induce actin stress fibers and focal adhesion sites in spreading human erythroleukemia (HEL) cells.

The effects of plasma fibronectin (pFn) and the tumor promoter 12-0-tetradecanoyl-phorbol-13-acetate (TPA) on adhesion and cytoskeletal organization of human erythroleukemia (HEL) cells were studied. HEL cells, that normally grow in suspension, attached rapidly on pFn-coated growth substratum and some cells showed spreading. Upon exposure to TPA most of the cells adhered and showed some degree of spreading also when plated on plastic. The spread cells showed mostly peripheral accumulations of F-actin in addition to actin fibers seen in some of the cells. When the cells were plated in the presence of TPA on pFn or on pFn-fragments, containing the cell binding site, all the cells adhered rapidly, spread extensively, organized prominent F-actin stress fibers and typical ventral plaques of vinculin and alpha-actinin. Both proteins were revealed also in the suspended cells by Western blot analysis. When plated on substratum coated with other pFn-fragments or laminin, the HEL cells did not adhere or spread. Both adhesion on pFn as well as formation of stress fibers in the presence of TPA could be prevented by the synthetic peptide Arg-Gly-Asp-Ser (RGDS). HEL cells were also able to organize typical ventral fibrillar arrays of Fn. Immunostaining and metabolic labeling experiments showed that the cells did not contain or synthesize Fn, indicating that the plaques were formed from exogenous pFn by the cells. The results suggest that Fn and TPA synergistically induce the organization of the actomyosin system in HEL cells by promoting the formation of prominent actin stress fibers and focal adhesion sites.

RGD peptides may only temporarily inhibit cell adhesion to fibronectin.

When human fibroblasts were cultured on fibronectin for 4 h in the presence of 0.5 mg/ml of the GRGDSP peptide derived from the fibronectin cell-binding site, they adhered and spread normally and organized talin and integrin alpha 5 and beta 1 subunits into focal adhesions. When the adherent cells were quantitated as a function of time, submaximal peptide concentrations were found to delay cell adhesion on fibronectin, but they had no effect on the maximum. When the cells were plated on vitronectin, however, even relatively low peptide concentrations lowered the maximal amount of cells adhering and abolished cell spreading. The results suggest a different mechanism for cell adhesion on fibronectin and vitronectin.

Involvement of the beta3 E749ATSTFTN756 region in stabilizing integrin alphaIIbbeta3-ligand interaction.

Platelet integrin alphaIIbbeta3 must be activated via intracellular mechanisms before it binds soluble ligands, and it is thought to be activated at its extracellular site by surface-bound ligands. Integrin activation is associated with rearrangement of the cytoskeleton and phosphorylation of proteins that become localized in focal contacts. In these processes, the cytoplasmic tail of the beta-subunit plays a central role. We introduced peptides homologous to the E749ATSTFTN756 domain (E-N peptide) and the T755NITYRGT762 domain (T-T peptide) of beta3 in streptolysin O-permeabilized platelets and analyzed the initial interaction with soluble fibronectin, fibrinogen and PAC-1 after stimulation with thrombin. E-N peptide left the initial binding of fibronectin intact but interfered with stable receptor occupancy. E-N peptide also inhibited fibrinogen binding, thereby reducing the formation of large aggregates. Strikingly, E-N peptide did not disturb the binding of PAC-1, which is known to reflect activation of the integrin. E-N peptide also inhibited tyrosine phosphorylation of focal adhesion kinase, a response known to be dependent on alphaIIbbeta3. T-T peptide did not affect these processes. In a model for outside-in integrin activation, E-N peptide disrupted the binding of CHO cells expressing alphaIIbbeta3 to surface-bound ligand. Again, T-T peptide had no effect. We conclude that the E749ATSTFTN756 region of the beta3-tail stabilizes the binding of soluble and surface-bound ligand to integrin alphaIIbbeta3 via a mechanism that involves the phosphorylation of FAK.

Conserved functions of the cytoplasmic domains of integrin beta subunits.

In this article I will discuss the notion that many integrins have similar functions in cell spreading, organization of the cytoskeleton and intracellular signalling. Most of these functions are transmitted through the cytoplasmic domains of integrin beta-subunits. These parts are also quite conserved between most integrins. I will discuss, what is known about the molecules binding to these parts of integrins, and which of those transmit the conserved functions.

Expression of integrins in human gingiva.

The distribution of the alpha 1-alpha 6 subunits of beta 1 integrins was studied by using a panel of monoclonal antibodies in indirect immunofluorescence microscopy. The results showed that the beta 1 subunit was expressed at the cell membrane of basal cells of gingival epithelium, throughout the cells of junctional epithelium (JE), and in cells of connective tissue, including endothelial cells and, more faintly, in inflammatory cells in gingival connective tissue. The alpha 4 subunit was expressed selectively in inflammatory cells, and the alpha 5 subunit was expressed in cells throughout gingival connective tissue. An overall cell membrane immunoreactivity for the alpha 2 and alpha 3 subunits was shown in basal cells of gingival epithelium and in cells of JE, corresponding to the epithelial localization of the beta 1 subunit. The alpha 6 subunit was polarized to the basal aspects of basal epithelial cells, but was also present in an overall cell surface distribution in basal cells and in cells of JE. The beta 4 integrin subunit was mainly expressed at the basal aspects of basal cells in gingival epithelium and JE. The results indicate that the alpha 2/beta 1, alpha 3/beta 1, alpha 6/beta 1, and alpha 6/beta 4 integrins are all expressed in human gingival epithelium. Of these, the alpha 6/beta 4 integrin complex is the major candidate for mediation of the attachment of epithelial cells to the basement membrane facing the connective tissue and probably also the tooth.

The Mr 140,000 fibronectin receptor complex in normal and virus-transformed human fibroblasts and in fibrosarcoma cells: identical localization and function.

We studied the function and localization of the fibronectin receptor complex in cultured normal and SV40-transformed human fibroblasts and in human fibrosarcoma cells by using monoclonal antibodies (MAbs) against the beta sub-unit of the receptor. Immunoprecipitation, fibronectin fragment affinity chromatography and immunoblotting results suggested that all the cells studied had similar amounts of the receptor. In normal fibroblasts MAbs additionally immunoprecipitated a smaller polypeptide, revealed as the precursor for the beta sub-unit and another polypeptide shown to be the alpha sub-unit of the VLA-I complex. The emergence of vinculin-positive focal adhesion sites and actin stress fibers was slower in the malignant cells than in normal fibroblasts when the cells were plated on non-coated glass-substrate in serum-free conditions and the fibronectin receptor complex did not become located to focal adhesions in any of the cells studied. Added substratum-bound but not soluble fibronectin mediated assembly of the fibronectin receptor complex to the focal adhesions in both normal and malignant cells. On fibronectin-coated growth substrate stress fibers also emerged as rapidly in the malignant cells as in normal fibroblasts. In all the cells the receptor complex, however, became largely dissociated from the focal adhesions within 48 hr. In cell adhesion conditions MAb against the alpha sub-unit of VLA-I complex revealed an even cell-surface labelling in normal fibroblasts and lack of labelling in malignant cells.

Human erythroleukemia cells adhere to fibronectin: evidence for a Mr 190,000-receptor protein.

Human erythroleukemia cells (K562) adhered rapidly on fibronectin (Fn)-coated growth substratum under serum-free conditions. The adhesion could be quantitatively inhibited by the synthetic peptide Arg-Gly-Asp-Ser (RGDS) and upon hemin-induced differentiation or trypsinization of the cells. Many of the cells also displayed rapid spreading that led to a redistribution of F-actin into spreading edges and in many cells also to a formation of typical actin fibers attaching to the ventral aspect of the cells. The spreading of the cells was inhibited by cytochalasin B but not by microtubule-disrupting drugs, suggesting an active role for the microfilament system in the spreading process. Direct overlay assay of electrophoretically separated polypeptides with 125I-Fn showed that in K562 cells there is a major Mr 190,000 Fn-binding protein that is lost upon differentiation. A similar overlay assay with purified plasma and cellular Fns followed by immunostaining with anti-Fn antibodies revealed a reaction with a similar polypeptide. The binding of Fns on the nitrocellulose sheets could be inhibited and the bound Fn eluted by using the RGDS peptide. From octylglucoside extracts of radioactively surface-labeled cells, distinct Mr 190,000/185,000 membrane glycoproteins bound to Fn-heptapeptide-Sepharose, further suggesting that the Mr 190,000 polypeptide would be the Fn-receptor of the K562 cells.

The effect of differentiation inducers on the integrin expression of K562 erythroleukemia cells.

We studied the effects of differentiation-inducers on the integrin profile and adhesive properties of K562 leukemia cells. The fibronectin (Fn) receptor integrin, alpha 5 beta 1, was the only integrin expressed in suspension cultured K562 cells. When the cells were exposed to 12-0-tetradecanoylphorbol-13-acetate (TPA) immunoreactivity for the beta 1 integrin subunit was slightly enhanced. TPA exposure also induced the appearance of the alpha 2, alpha 3, alpha v and beta 3 integrin subunits, but the platelet integrin subunit alpha IIb was not detected. On the other hand, hemin chloride-induced erythroid differentiation of K562 cells diminished the expression of the alpha 5 beta 1 integrin on the surface of the cells. Adhesion experiments with TPA-exposed K562 cells indicated that although the adherence to the extracellular matrix (ECM) proteins as a rule was low a few cells spread on these proteins. The present results specify the effects of differentiation inducers on the integrin profile of K562 cells and excludes the comprehension that TPA would induce expression of the platelet integrin alpha IIb on their surface. Our results also show, that an increased expression of a certain integrin does not necessarily lead to a comparable adhesion ability on its ligand in vitro.

Effects of mutations in the cytoplasmic domain of integrin beta(1) to talin binding and cell spreading.

Integrins are transmembrane proteins linking the extracellular matrix or certain cell-cell contacts to the cytoskeleton. To study integrin-cytoskeleton interactions we wanted to relate talin-integrin interaction to integrin function in cell spreading and formation of focal adhesions. For talin-binding studies we used fusion proteins of glutathione S-transferase and the cytoplasmic domain of integrin beta(1) (GST-cytobeta(1)) expressed in bacteria. For functional studies chimeric integrins containing the extracellular and transmembrane parts of beta(3) linked to the cytoplasmic domain of beta(1) were expressed in CHO cells as a dimer with the alpha(IIb) subunit. Point mutations in the amino acid sequence N(785)PIY(788) of beta(1) disrupted both the integrin-talin interaction and the ability of the integrin to mediate cell spreading. COOH-terminal truncation of beta(1) at the amino acid position 797 disrupted its ability to mediate cell spreading, whereas the disruption of talin binding required deletion of five more amino acids (truncation at position 792). A synthetic peptide from this region of beta(1) (W(780)DTGENPIYKSAV(792)) bound to purified talin and inhibited talin binding to GST-cytobeta(1). The ability of the mutants to mediate focal adhesion formation or to codistribute to focal adhesions formed by other integrins correlated with their ability to mediate cell spreading. These results confirm the previous finding that a talin-binding site in the integrin beta(1) tail resides at or close to the central NPXY motif and suggest that the integrin-talin interaction is necessary but not sufficient for integrin-mediated cell spreading.

UNC-87 is an actin-bundling protein.

The Caenorhabditis elegans unc-87 gene product is essential for the maintenance of the nematode body wall muscle where it is found colocalized with actin in the I band. The molecular domain structure of the protein reveals similarity to the C-terminal repeat region of the smooth muscle actin-binding protein calponin. In this study we investigated the in vitro function of UNC-87 using both the full-length recombinant molecule and several truncated mutants. According to analytical ultracentrifugation UNC-87 occurs as a monomer in solution. UNC-87 cosedimented with both smooth and skeletal muscle F-actin, but not with monomeric G-actin, and exhibited potent actin filament bundling activity. Actin binding was independent of the presence of tropomyosin and the actin cross-linking proteins filamin and alpha-actinin. Consistent with its actin bundling activity in vitro, UNC-87 tagged with green fluorescent protein associated with and promoted the formation of actin stress fiber bundles in living cells. These data identify UNC-87 as an actin-bundling protein and highlight the calponin-like repeats as a novel actin-binding module.

Regulation of integrin affinity states through an NPXY motif in the beta subunit cytoplasmic domain.

The ligand binding affinities of the integrins are regulated through their cytoplasmic domains. To identify specific residues that are involved in this process, we have generated mutants in the beta 1 and beta 3 tails and coexpressed them in Chinese hamster ovary cells with constitutively active alpha subunits. These alpha subunits are chimera of extra-cellular and transmembrane alpha IIb joined to the cytoplasmic domains of alpha 5, alpha 6A, or alpha 6B and confer an energy-dependent high affinity state when expressed in Chinese hamster ovary cells. The affinity state of these transfectants was determined by analyzing the binding of PAC1, an antibody that specifically recognizes the activated form of the reporter group, extracellular alpha IIb beta 3. We have identified point mutants in several areas of the beta tails, which result in a reduced ability to bind ligand. Complete abolition of PAC1 binding was obtained with mutants in an NPXY motif found in many integrin beta subunits and implicated in the internalization of other cell surface receptors. Similar effects on PAC1 binding were observed whether coexpression was with alpha chimera containing alpha 5, alpha 6A, or alpha 6B cytoplasmic sequences. These studies identify a novel role for the NPXY motif in the regulation of integrin binding affinity.

Human natural killer cells express different integrins and spread on fibronectin.

Human natural killer (NK) cells adhered and most of them also actively spread on cellular fibronectin (cFn), plasma Fn (pFn) and its Mr 120,000-140,000 or Mr 105,000 cell-binding proteolytic Fn-fragments as well as on heparin-binding Fn-fragments containing the alternative cell binding site. The cells did not spread on vitronectin, laminin or collagens. Adhesion on Mr 105,000 Fn fragment containing the cell binding site, could be prevented by the synthetic peptide GRGDS but not by an inactive peptide, whereas adhesion on heparin-binding Fn fragments was unaffected by the peptide. Spreading of the NK cells led to a distinct reorganization of F-actin. Immunoprecipitation with monoclonal antibodies (MoAb) against the beta 1 integrin subunit of radioactively surface-labelled cells revealed a broad polypeptide band of Mr 140,000 under reducing conditions and a polypeptide doublet of Mr 160,000 and Mr 110,000 under non-reducing conditions. Identical polypeptides, corresponding to the alpha- and beta-subunits of the Fn-receptor complex, were bound to the Mr 105,000 chymotryptic Fn-fragment coupled to Sepharose. Similar experiments with small lymphocytes did not reveal any polypeptides. Immunofluorescence results with McAbs suggested that among the alpha-subunits of integrins, the alpha 3, alpha 4, and alpha 5 subunits are expressed in NK cells. The present results suggest that non-activated NK cells, but not small lymphocytes, express beta 1-integrins, and that at least the Fn-receptors alpha 4 beta 1 and alpha 5 beta 1 may function in the adhesion and migration of NK cells.

Distribution of beta 1 and beta 3 integrins in human fetal and adult kidney.

The distribution of beta 1 and beta 3 integrins was studied in fetal and adult human kidneys by indirect immunofluorescence microscopy. In the developing kidney, the cells of the undifferentiated metanephric blastema displayed strong cell surface-confined beta 1 integrin immunoreactivity, whereas the cells of primary vesicles and comma- and S-shaped bodies reacted more weakly. In mature fetal as well as adult glomeruli, beta 1 integrins were distinctly localized, apparently confining to the basal cell surfaces of endothelial cells and podocytes abutting the glomerular basement membrane. In adult proximal tubules, beta 1 integrin immunoreactivity was strictly confined to the basal aspect of the epithelial cells, being absent laterally, which is unusual for membrane proteins of polarized epithelial cells. A more diffuse overall immunoreactivity was seen in distal tubules and collecting ducts. The epithelial cells of developing proximal and distal tubules displayed an overall distribution of beta 1 integrins. In each case, talin immunoreactivity followed that of beta 1 integrins. Compared with beta 1 integrins, beta 3 integrins showed a more restricted distribution, and differences were seen in the reactions of mono- and polyclonal antibodies. In developing glomeruli, beta 3 integrin immunoreactivity was prominently seen in the cells of Bowman's capsule, possibly revealing the presence of vitronectin receptor. Solitary cells, that reacted also with antibodies to the platelet glycoprotein IIb, were consistently detected in fetal glomeruli, suggesting the presence of megakaryocytes. The results show that during nephrogenesis, beta 1 integrins become distinctly polarized both in glomerular endothelial cells and podocytes, as well as in the epithelial cells of proximal tubules.

Localization of beta 1, beta 3, alpha 5, alpha v, and alpha IIb subunits of the integrin family in spreading human erythroleukemia cells.

The localization of five integrin subunit proteins was studied in human erythroleukemia (HEL) cells spreading on various culture substrata in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) and the absence of serum. The cells readily adhered on fibronectin, but TPA was needed for adherence on vitronectin and for the spreading of the cells on both substrata. Indirect immunofluorescence microscopy showed that in the spread cells cultured on vitronectin or fibronectin for 2 hours, beta 1, beta 3, alpha 5, and alpha IIb integrin subunits were localized at focal adhesions as identified by talin-immunoreactivity. The alpha v integrin immunoreactivity was initially found at the focal adhesions when the cells were cultured on vitronectin, but was also found later in cells cultured on fibronectin. The alpha IIb integrin immunoreactivity disappeared from focal adhesions within 24 hours. The alpha 5 and beta 1 integrin immunoreactivities disappeared from the focal adhesions in cells cultured on vitronectin, but not in cells cultured on fibronectin. When the cells were plated on glass substratum in the presence of TPA, they spread much slower than on vitronectin or fibronectin, but some cells showed focal adhesions after only 8 hours in culture. In this case, the alpha v and beta 3 integrin subunits were found at focal adhesions. After TPA treatment, HEL cells deposited thrombospondin-immunoreactive material onto their culture substratum, but synthesis of fibronectin, vitronectin, fibrinogen, or von Willebrand factor was not detected. Thus, the results suggest that TPA would activate several integrin receptors in HEL cells and also stimulate the secretion of thrombospondin, which might be used as an adhesion ligand for the integrin vitronectin receptor alpha v/beta 3 complex.

Distribution of the alpha 1-alpha 6 integrin subunits in human developing and term placenta.

The distribution of the alpha 1-alpha 6 as well as alpha v, beta 1, beta 3 and beta 4 integrin subunits in human first and second trimester and term placentas was studied by indirect immunofluorescence microscopy using a panel of monoclonal antibodies (mAbs). In first and second trimester villi, the alpha 1 and beta 1 integrin subunits were detected in the stromal cells, that were mostly also immunoreactive for desmin. Desmin-positive stromal cells were also found in villi of term placentas, but the stroma was negative for anti-alpha 1 and -beta 1. In the villous trophoblast, anti-alpha 6 and -beta 4 revealed a distinct basal immunoreactivity during all stages of development, whereas immunoreactivity for the alpha 3 and beta 1 subunits emerged during the second and third trimesters. Throughout placental development, endothelia of villous capillaries reacted prominently with anti-alpha 1 and -beta 1. Intermediate trophoblastic cells displayed a somewhat heterogenous immunoreactivity for the beta 1, alpha 1, alpha 3 and alpha 5 integrin subunits, and differed from villous trophoblast also in their lack of expression of the alpha 6 and beta 4 subunits. While nondecidualized endometrial cells displayed weak reactivity for the alpha 1 and beta 1 integrin subunits, the individual decidual cells presented both a basement membrane and a cell surface-confined immunoreactivity for anti-alpha 1, -alpha 3, and -beta 1. The results suggest a role for integrins in placental development, and show that expression of integrins is modulated during the differentiation of trophoblast, villous stroma, and decidual cells. Furthermore, the basal localization of alpha 6 beta 4 and alpha 3 beta 1 integrins suggests that they are employed as basement membrane receptors in the villous trophoblast, and the emergence of the alpha 3 beta 1 complex may reflect that the cytotrophoblast and syncytiotrophoblast recognize the basement membrane differently.