The influence of neuropeptides related to pro-opiomelanocortin on acquisition of heroin self-administration of rats

The influence of different neuropeptides related to pro-opiomelanocortin were tested on acquisition of heroin self-administration in rats. The animals were allowed to self-administer heroin intravenously on a continuous reinforcement schedule during 6 h daily sessions on 5 consecutive days. Treatment was performed subcutaneously 1 h before each daily session. It was found that the opioid peptides alpha-, gamma- and beta-endorphin hardly influenced acquisition of heroin self-administration, while the non-opioid fragments of alpha- and gamma- endorphin modulated this behavioral response. In fact, beta-endorphin (beta E) 2-9 tended to facilitate the rate of acquisition, while the gamma-type endorphins, des-Tyr1-gamma-endorphin (beta E 2-17) and des-enkephalin-gamma-endorphin (beta E 6-17), decreased heroin intake. Concerning the ACTH/MSH related peptides, a decreasing effect of heroin intake was found following treatment with (D-Phe7)-ACTH 4-10, with a high dose of the ACTH 4-9 analog Org 2766 and with gamma 2-MSH, while ACTH 1-24, ACTH 4-10 and a low dose of Org 2766 did not significantly influence self-injecting behavior. It is concluded that pro-opiomelanocortin serves as a precursor molecule for peptide fragments, which modulate the acquisition of heroin self-administration in rats.     

Differential in vitro metabolism of beta-endorphin in schizophrenia

Biologically active peptide fragments derived from the proteolytic cleavage of beta-endorphin (beta E) have been shown to be present in the brain. Based on clinical results using some of these fragments in neuropsychiatric disease studies we investigated the in vitro metabolism of beta E by twice-washed membrane homogenates of postmortem putamen from sex and age matched controls versus subjects with a diagnosis of schizophrenia. The present study demonstrates that frozen (-80 degrees C) postmortem human tissues are viable for these studies and that metabolism in control tissue proceeds similarly to fresh tissues. Furthermore, a significant increase in the formation of the putative neuroleptic-like peptide fragment des-enkephalin-gamma-endorphin in postmortem schizophrenic putamen versus controls was shown. A significant decrease in the formation of beta E was also reported. These data suggest that an approach using postmortem human brain is possible in studying beta-endorphin catabolism and is therefore applicable to other neuropeptide systems.     

Interaction of human beta-endorphin with nonopiate binding sites on the terminal SC5b-9 complex of human complement. Significance of COOH-terminal beta H-endorphin fragments

We have characterized the binding of 125I-labeled human beta-endorphin (125I-beta H-endorphin) to sites present on the terminal fluid-phase complex of human complement, consisting of complement components C5b, C6, C7, C8, C9, and the S-protein (SC5b-9 complex). Specific binding exhibited saturability, reversibility, structural specificity, temperature dependence, and absence of negative cooperative effects. Binding was maximal at 4 degrees C and pH 7.0; it was diminished by monovalent and divalent cations as well as by increasing concentrations of urea and Triton X-100 and apparently required intact disulfide groups. Binding was not inhibited by a number of opioid peptides sharing common sequences with the NH2 terminus of beta H-endorphin. In contrast, binding was inhibited by beta H-endorphin, N-acetyl-beta H-endorphin, and a series of COOH-terminal beta H-endorphin fragments, where of the COOH-terminal dipeptide Gly-Glu represented the minimal effective structure. Stepwise extension towards the NH2 terminus led to an increased binding affinity of the respective fragment. Computer resolution of competition curves yielded one binding component for several shorter COOH-terminal beta H-endorphin fragments and for beta H-endorphin (1-5) + (16-31), whereas two distinct binding components were obtained when beta H-endorphin (27-31), beta H-endorphin (6-31), N-acetyl-beta H-endorphin or beta H-endorphin were used as inhibitors. This study presents detailed data on the binding of COOH-terminal beta H-endorphin fragments to specific nonopiate binding sites present on the terminal SC5b-9 complex of human complement. We suggest that through this interaction, beta H-endorphin may modulate certain functions within the immune system.     

Beta(Leu121-Lys122) segment of fibrinogen is in a region essential for plasminogen binding by fibrin fragment E

It was shown previously that two sequentially nonidentical regions of human fibrin(ogen), present in fragments D and E, carry specific plasminogen-binding sites [V aradi , A., & Patthy , L. (1983) Biochemistry 22, 2440-2446]. Comparison of the affinity of a variety of fragment E species for immobilized Lys-plasminogen revealed that fragment E3e [(alpha 20/24-78, beta 54-122, gamma 1-53)2] possesses a strong plasminogen-binding site, whereas fragment E3t [(alpha 20/24-78, beta 54-120, gamma 1-53)2] has 30-fold lower affinity for the affinant . Since the two fragments differ only in the beta ( Leu121 - Lys122 ) segment, this suggests that residues beta ( Leu121 - Lys122 ), present in the triple-helical connector region of fibrin(ogen), are essential for plasminogen binding by fragment E. Reduction and alkylation of fragment E3e lead to the destruction of the plasminogen-binding site, indicating that none of the separated, alkylated polypeptide chains of the fragment are able to bind to plasminogen and probably the coiled-coil superstructure of the connector region is necessary for the maintenance of the plasminogen-binding site of fragment E.     

Interaction of fibrin(ogen) with heparin: further characterization and localization of the heparin-binding site

The beta chain 15-42 sequence of the fibrin(ogen) E region was implicated in heparin binding [Odrljin et al. (1996) Blood 88, 2050-2061]; whether heparin binds to other fibrin(ogen) regions remains to be clarified. To address this question, we studied the interaction of heparin with fibrinogen, fibrin, and their major fragments D(1), D-D, E(1), E(3), and alphaC, which together cover the entire structure of the molecule, by ligand blotting, surface plasmon resonance, and fluorescence. All three techniques revealed that at physiological ionic conditions only fibrin(ogen) and the E(1) fragment bind heparin, indicating that the only physiologically relevant heparin-binding site of fibrin(ogen) is located in its E region. To test whether the beta15-42 sequence is sufficient to form this site or some additional sequences are also involved, we tested the interaction of heparin with a number of beta15-42-containing fragments. The synthetic beta15-42 peptide bound heparin weakly (K(d) = 44.5 microM) while the recombinant beta15-57 and beta15-64 fragments exhibited almost 7-fold higher affinity (K(d) = 6.4 and 7.1 microM, respectively), indicating that the beta43-57 region is also important for heparin binding. At the same time the recombinant dimeric disulfide-linked (beta15-66)(2) fragment which mimics the dimeric arrangement of the beta chains in fibrin bound heparin with high affinity (K(d) = 66 nM), almost 100-fold higher than that for the monomeric fragments. This affinity was similar to those determined for fibrin and the E(1) fragment (K(d) = 72 and 70 nM, respectively) suggesting that (beta15-66)(2) mimics well the heparin-binding properties of the latter two. Altogether, these results indicate that the only heparin-binding site in fibrin(ogen) is formed by NH(2)-terminal portions of the beta chains, including residues beta15-57, and that dimerization is essential for high-affinity binding.     

The alpha 1/beta 1 and alpha 6/beta 1 integrin heterodimers mediate cell attachment to distinct sites on laminin

This study was undertaken to determine the roles of individual alpha/beta 1 integrin heterodimers in promoting cellular interactions with the different attachment-promoting domains of laminin (LN). To do this, antibodies to the integrin beta 1 subunit or to specific integrin alpha subunits were tested for effects on cell attachment to LN, to elastase fragments E1-4 and E1, derived from the short arms and core of LN's cruciform structure, and to fragment E8 derived from the long arm of this structure. The human JAR choriocarcinoma cells used in this study attached to LN and to fragments E1 and E8. Attachment to E1-4 required a much higher substrate coating concentration, suggesting that it is a poor substrate for JAR cell attachment. The ability of cells to attach to different LN domains suggested the presence of more than one LN receptor. These multiple LN receptors were shown to be beta 1 integrin heterodimers because antibodies to the integrin beta 1 subunit inhibited attachment of JAR cells to LN and its three fragments. To identify the individual integrin alpha/beta 1 heterodimers that mediate interactions with these LN domains, mAbs specific for individual beta 1 heterodimers in human cells were used to study JAR cell interactions with LN and its fragments. An anti-alpha 6/beta 1-specific mAb, GoH3, virtually eliminated cell attachment to E8 and partially inhibited attachment to E1 and intact LN. Thus the major alpha 6/beta 1 attachment domain is present in fragment E8. An alpha 1/beta 1-specific mAb (S2G3) strongly inhibited cell attachment to collagen IV and partially inhibited JAR attachment to LN fragment E1. Thus, the alpha 1/beta 1 heterodimer is a dual receptor for collagen IV and LN, interacting with LN at a site in fragment E1. In combination, the anti-alpha 1- and anti-alpha 6-specific antibodies completely inhibited JAR cell attachment to LN and fragment E1. Thus, the alpha 1/beta 1 and alpha 6/beta 1 integrin heterodimers each function as LN receptors and act together to mediate the interactions of human JAR choriocarcinoma cells with LN.     

Analysis of the beta-endorphin structure-related activity on human monocyte chemotaxis: importance of the N- and C-terminal

We evaluated the chemotactic activity of beta-endorphin and beta-endorphin-related peptides on human monocytes. We tested beta-endorphin(1-31) and fragments (1-16), (1-17), (1-27) in which the N-terminal of the opioid is preserved, N-acetyl-beta-endorphin(1-31) and fragments (6-31) and (28-31) in which the C-terminal is preserved, and fragment (2-17) that lacks both the N- and C-terminal. The fragments in which the N- and C-terminal were preserved [with the exception of fragment (28-31)] showed a chemotactic effect, while the lack of both terminals deprived the peptides of any activity. Moreover, only the N-terminal-mediated effects were naloxone reversible, while the C-terminal effects were not. These results indicate that while the intact N-terminal is necessary for opioid like effects, both N- and C-terminal can mediate effects on the immune system, thus offering evidence for a nonopioid receptor-mediated effect of opioid peptides on the immune system.     

Structure of fragment E species from human cross-linked fibrin

Fragments E1, E2, and E3 are plasmic derivatives of fibrin encompassing the NH2-terminal region of the molecule. The first two species, but not the third, can bind to fragment DD, forming a (DD)E complex, and therefore probably contain binding sites involved in the polymerization of fibrin. For localization of these sites the structure of the fragments was determined by establishing the NH2- and COOH-terminal boundaries of the molecules and using the published amino acid sequence of fibrinogen. Fragment E1 encompasses Gly-alpha 17 to Lys-alpha 78, Gly-beta 15 to Lys-beta 122, and Tyr-gamma 1 to Lys-gamma 62, this representing the intact NH2-terminal region of fibrin. Fragment E2 is an asymmetric molecule which is lacking the sequence of Gly-beta 15 to Lys-beta 53 in one beta-chain remnant. This fragment E2 also lost Lys-beta 122 from the COOH terminal of the beta chain as compared with fragment E1. These cleavages did not affect the ability of fragment E2 to bind to fragment DD. Fragment E3 was heterogeneous, the main species encompassing Val-alpha 20 to Lys-alpha 78, Lys-beta 54 to Leu-beta 120, and Tyr-gamma 1 to Lys-gamma 53. Thus, the loss of the binding function involved in the formation of fibrin clot was associated with the removal of small fragments from all three polypeptide chains: alpha 17-19 (Gly-Pro-Arg), beta 15-53 from the remaining half of the molecule, beta 121 (Leu), and gamma 54-58 (Thr-Ser-Glu-Val-Lys).     

des-Tyr1-gamma-endorphin and haloperidol increase pineal gland melatonin levels in rats

The effect of subcutaneously injected DT gamma E (beta-endorphin, (beta E)2-17) on the pineal melatonin level was compared with that of closely related peptides and the neuroleptic drug haloperidol. As found previously, DT gamma E (3 ng/rat and 300 ng/rat) increased the melatonin levels. Similar doses of DT alpha E (beta E 2-16), DT beta E (beta E 2-31), gamma E (beta E 1-17), alpha E (beta E 1-16) and beta E failed to significantly change the melatonin levels in both the dark and the light phase. Haloperidol in a dose of 300 ng/rat exhibited a similar effect as DT gamma E.     

Skeletal myoblasts utilize a novel beta 1-series integrin and not alpha 6 beta 1 for binding to the E8 and T8 fragments of laminin.

The E8 fragment of laminin stimulates myoblast attachment and locomotion. Myoblast attachment to laminin/E8 was blocked by anti-integrin antibodies against beta 1-chains but not by antibodies against alpha 6-chains. By contrast, other cell lines (e.g. B16, HT1080, P19, F9, Pys2, 3T3, and 3T6) were blocked both by anti-beta 1 and anti-alpha 6. All cells tested also bound to approximately 125-kDa C-terminal fragments of E8 (T8 and T8'). Immunoprecipitation of surface-iodinated myoblasts revealed beta 1-, alpha 3-, and alpha 5-integrin chains and a novel chain that co-precipitated with anti-beta 1 antibodies running at approximately 95 kDa (reduced). I125-alpha 6 beta 1 was immunoprecipitated from cells whose attachment to E8 was blocked by anti-alpha 6 antibodies. By contrast, little alpha 6 beta 1 could be immunoprecipitated from myoblasts. beta 1-Integrin and the novel alpha-chain (alpha'), Mr approximately 120,000/approximately 95.000 (nonreduced/reduced), from myoblast lysates were retained during affinity chromatography on Engelbreth-Holm-Swarm-laminin affinity columns. beta 1, alpha 1, and the novel alpha' were retained from Rugli cell lysates on Engelbreth-Holm-Swarm-laminin columns. alpha 3 was not bound. When E8 was used as affinity matrix, only beta 1 and alpha' were retained. The N-terminal sequence of Rugli alpha' was homologous to alpha-chains of beta 1-series integrins and was most similar to alpha 6 (9 identical residues out of 14). However, there were distinctive differences; in particular, 2 residues were deleted in comparison with alpha 6.     

Characterization of beta-endorphin in human pituitary by fast atom bombardment mass spectrometry of trypsin-generated fragments

A novel mass spectrometric method possessing a high level of structural specificity is described for characterization in biological fluids and tissues of endogenous beta-endorphin of the human amino acid sequence (beta h-EP). The method is based upon purification of tissue extracts by an RP-HPLC gradient, followed by trypsinolysis of that particular HPLC fraction corresponding to the elution time of synthetic beta h-EP. The tryptic digest of that endogenous beta h-EP fraction was purified further by a second RP-HPLC gradient. A unique tryptic fragment selected from the second gradient was analyzed by fast atom bombardment mass spectrometry and B/E linked-field scan MS/MS techniques to provide molecular weight and amino acid sequence-determining fragment ion information, respectively, of that fragment. Collectively, these independent analytical methodologies provided unequivocal structure evidence for the presence of endogenous beta h-EP in human pituitary. The method was established first by utilizing synthetic beta h-EP to optimize experimental parameters, and then applied to the analysis of beta h-EP in post-mortem human pituitary extracts. The suitability of the present method for semi-quantitation of tissue extracts is also demonstrated. The corresponding detection limit of the synthetic beta h-EP was 90 fmol, and human pituitary contained 1.5 pmol of beta h-EP mg-1 protein. The method can be extended readily to the analysis of beta-endorphin derived from other species and tissues.     

HU and IHF, two homologous histone-like proteins of Escherichia coli, form different protein-DNA complexes with short DNA fragments.

Using the gel retardation technique we have studied the protein-DNA complexes formed between HU--the major histone-like protein of Escherichia coli--and short DNA fragments. We show that several HU heterodimers bind DNA in a regularly spaced fashion with each heterodimer occupying about 9 base pairs. The alpha 2 and beta 2 HU homodimers form the same structure as the alpha beta heterodimer on double stranded DNA. However when compared to the heterodimer, they bind single stranded DNA with higher affinity. We also show that HU and the Integration Host Factor of E. coli (IHF) form different structures with the same DNA fragments. Moreover, HU seems to enhance the DNA-binding capacity of IHF to a DNA fragment which does not contain its consensus sequence.     

The peptide beta 43-47 which increases microvascular permeability is released by plasmin during cleavage of fragment Y of fibrinogen

A synthetic pentapeptide corresponding to sequence 43-47 of human fibrinogen B beta chain elicited, in rabbits, antibodies that during immunoblotting recognized intact fibrinogen, fragments X and Y as well as the B beta chain. Since fragment Y is the last peptide product which reacts with anti-beta 43-47 antibodies, splitting of fragment Y into fragment D and fragment E must be accompanied by plasmin cleavage of the peptide bond beta Lys-47-Ala-48.     

Dependence of RGS9-1 membrane attachment on its C-terminal tail.

RGS9-1 is a GTPase-accelerating protein (GAP) required for rapid recovery of the light response in vertebrate rod and cone photoreceptors. Similar to its phototransduction partners transducin (G(t)) and cGMP phosphodiesterase, it is a peripheral protein of the disc membranes, but it binds membranes much more tightly. It lacks the lipid modifications found on G(t) and cGMP phosphodiesterase, and the mechanism for membrane attachment is unknown. We have used limited proteolysis to generate a fragment of RGS9-1 that is readily removed from membranes under moderate salt conditions. Immunoblots reveal that this soluble fragment lacks a 3-kDa fragment from the C-terminal domain, the only domain within RGS9-1 that differs in sequence from the brain-specific isoform RGS9-2. Recombinant fragments of RGS9-1 with or without the partner subunit G beta(5L) were constructed with or without the C-terminal domain. Those lacking the C-terminal domain bound to photoreceptor membranes much less tightly than those containing it. Removal by urea of G beta(5L) from endogenous or recombinant RGS9-1 bound to rod outer segment membranes left RGS9-1 tightly membrane-bound, and recombinant RGS9-1 was urea-soluble in the absence of membranes. Thus the C-terminal domain of RGS9-1 is critical for membrane binding, whereas G beta(5L) does not play an important role in membrane attachment.     

Binding of beta-endorphin and its fragments to the non-opiod receptor of murine peritoneal macrophages

The tritium-labeled selective agonist of the nonopioid beta-endorphin receptor the decapeptide immunorphin ([3H]SLTCLVKGFY) with a specific activity of 24 Ci/mmol was prepared. It was shown that [3H]immunorphin binds with a high affinity to the non-opioid beta-endorphin receptor of mouse peritoneal macrophages (Kd 2.4 +/- 0.1 nM). The specific binding of [3H]immunorphin to macrophages was inhibited by unlabeled beta-endorphin (Ki of the [3H]immunorphin-receptor complex 2.9 +/- 0.2 nM) and was not inhibited by unlabeled naloxone, alpha-endorphin, gamma-endorphin, and [Met5]enkephalin (Ki > 10 microM). Thirty fragments of beta-endorphin were synthesized, and their ability to inhibit the specific binding of [3H]immunorphin to macrophages was studied. It was found that the shortest peptide having practically the same inhibitory activity as beta-endorphin is its fragment 12-19 (Ki 3.1 +/- 0.3 nM).     

Beta-endorphin and related peptides suppress phorbol myristate acetate-induced respiratory burst in human polymorphonuclear leukocytes

In the present study, the immunomodulatory effect of beta-endorphin (beta-E) and shorter pro-opiomelanocortin (POMC) fragments was evaluated by assessing their influence on respiratory burst in human polymorphonuclear leukocytes (PMN). The effect of the peptides (10(-17)M - 10(-10)M) on phorbol myristate acetate (PMA)-stimulated production of reactive oxygen metabolites was measured in a lucigenin-enhanced chemiluminescence (CL) assay. Both POMC peptides with opiate-like activity (i.e. alpha-endorphin (alpha-E), beta-E and gamma-endorphin (gamma-E] and their non-opioid derivatives (i.e. des-TYR1-beta-endorphin (dT beta E), des-TYR1-gamma-endorphin (dT gamma E), and des-ENK-gamma-endorphin (dE gamma E] were tested. With the exception of alpha-E, PMA-stimulated respiratory burst was suppressed by all POMC fragments tested. A U-shaped dose-response relation was observed. Doses lower than 10(-17)M and higher than 10(-8)M were without effect. beta-E and dT beta E both suppressed PMA-induced oxidative burst in human PMN at physiological concentrations (10(-16)M - 10(-10)M). gamma-E and dT gamma E proved to be less potent inhibitors, reaching maximal effect at higher concentrations (10(-12)M - 10(-10)M). DE gamma E exerted an even less pronounced but still significant suppressive effect at the concentration of 10(-10)M. None of the endorphins tested was shown to affect resting oxidative metabolism in the PMN. The modulatory effects of the opioid peptides could not be blocked by the opioid antagonist naloxone (10(-8)M). These data show that fragments derived from the POMC-precursor molecule modulate the activation of PMN by suppressing PMA-stimulated oxidative metabolism and that this activity does not involve a classical opiate-like receptor.     

Binding characteristics of dermorphin, and [dermorphin1-7]-beta c-endorphin in rat brain membranes

Dermorphin and a camel beta-endorphin (beta c-EP) analog in which residues 1-7 correspond to the dermorphin sequence ([Dermorphin1-7]-beta c-EP) have been investigated with respect to their receptor binding characteristics using human and camel beta-EP as reference peptides. Tritiated dihydromorphine, [D-Ala2, D-Leu5]-enkephalin, ethylketocyclazocine and human beta-endorphin were used as primary ligands in the rat brain membrane preparation for radioreceptor assay. Camel beta-endorphin was the most potent peptide in all experiments. [Dermorphin1-7]-beta c-EP is significantly less potent towards 3H-ethylketocyclazocine and 3H-[D-Ala2, D-Leu5]-enkephalin but is as potent towards 3H-dihydromorphine and 3H-human beta-endorphin. Dermorphin itself weakly displaces tritiated dihydromorphine, [D-Ala2, D-Leu5]-enkephalin and ethylketocyclazocine (potency relative to camel beta-EP, 1-4%) but it is more potent (9%) in competition with tritiated human beta-endorphin. Dermorphin and the [Dermorphin-1-7]-beta c-EP appear to interact preferentially with mu opiate receptors.     

Human beta-endorphin. Synthesis and biological activity of analogs with modification in residue positions 8, 27, and 9 or 11

Three analogs of human beta-endorphin (beta h-ER) were synthesized by the solid-phase method: [Gln8,Trp27]-beta h-EP (I), [Gln8,Arg9,Trp27]-beta h-EP (II), and [Gln9,Arg11,Trp27]-beta h-EP (III). Radioreceptor binding assay with use of tritiated beta h-EP as primary ligand gave relative potencies as follows: beta h-EP, 100;I, 778;II, 467;III, 449. Relative potencies in an analgesic assay were: beta h-EP, 100;I, 114;II, 165;III, 83. The 8-11 segment of beta h-EP can tolerate a net increase in charge of +2 without diminishing analgesic potency. The substitution of Glu8 may be one of the more dependable means of designing beta-endorphin antagonists.     

Integrin recognition of different cell-binding fragments of laminin (P1, E3, E8) and evidence that alpha 6 beta 1 but not alpha 6 beta 4 functions as a major receptor for fragment E8

The involvement of integrins in mediating interaction of cells to well-characterized proteolytic fragments (P1, E3, and E8) of laminin was assessed by antibody blocking studies. Cell adhesion to fragment P1 was affected by mAbs against the integrin beta 1 and beta 3 subunits and furthermore could be prevented completely by a synthetic peptide containing the Arg-Gly-Asp sequence. Because the beta 3 antibody-sensitive cell lines expressed the vitronectin receptor (alpha v beta 3) at high levels, the involvement of this receptor in cell adhesion to P1 is strongly suggested. Integrin-mediated cell adhesion to E3 is of low affinity and was inhibited by antibodies against the integrin beta 1 subunit. In contrast, adhesion of some cell types to E3 was not or only partially sensitive to inhibition by anti-integrin subunit antibodies. Cell adhesion to E8 was blocked completed by integrin alpha 6 or beta 1 antibodies. The alpha 6-specific antibody did not inhibit cell adhesion to E3 or P1. Furthermore, the antibody only blocked adhesion to laminin of those cells that adhered exclusively to the E8 fragment. In addition, expression of alpha 6 beta 1 was closely correlated with the ability of cells to bind to the E8 fragment of laminin. These results indicate that the alpha 6 beta 1 integrin is a specific receptor for the E8 fragment of laminin. Many cell types expressed, instead of or in addition to alpha 6 beta 1 the recently described integrin alpha 6 beta 4. Although the ligand of alpha 6 beta 4 was not identified, it must be different from that of alpha 6 beta 1, because cells that express alpha 6 beta 4, but not alpha 6 beta 1, do not adhere to E8, and cell adhesion to E8 was specifically blocked by beta 1 specific antibodies. In conclusion, the data indicate that distinct integrin receptors belonging to the beta 1 or beta 3 subfamily are involved in adhesion of cells to the various laminin fragments. Adhesion to E3 may also be brought about by other receptor molecules, possibly proteoglycans, not belonging to the integrin family.