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Results for “functional divergence”

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At least 109 records · Page 6Linked to original sources
PubMed1994

Integrin alpha v beta 8. Interaction with vitronectin and functional divergence of the beta 8 cytoplasmic domain.

The integrin beta 8 subunit was identified by cloning and sequencing of the cDNA and has been shown to associate with the alpha v subunit (Moyle, M., Napier, M. A., and McLean, J. W. (1991) J. Biol. Chem. 266, 19650-19658). We now present initial data on its functional properties. We produced a recombinant secreted form of alpha v beta 8 and used it to raise monoclonal antibodies that recognize the alpha v beta 8 complex or the beta 8 subunit alone on the surface of melanoma cells and on beta 8-transfected human embryonic kidney (293) cells. Affinity chromatography experiments showed that secreted alpha v beta 8 bound to vitronectin but not to fibronectin, collagen, or fibrinogen. Supporting evidence that intact full-length alpha v beta 8 could also bind to vitronectin-Sepharose was provided by performing affinity chromatography with the melanoma cell line MeWo, which normally expresses the intact beta 8 subunit. By studying the adhesive properties of melanoma cells and beta 8-transfected 293 cells, we found that alpha v beta 8 by itself does not promote cell adhesion on a vitronectin-coated substrate. To test the respective functional activities of the beta 8 extracellular and cytoplasmic domains, we analyzed chimeric beta 8/beta 3 subunit constructs. The beta 3 subunit was chosen because full-length beta 3, when transfected into 293 cells, strongly supports cell adhesion. We found that a chimeric integrin containing the beta 3 extracellular domain combined with the beta 8 transmembrane and cytoplasmic domains did not promote 293 cell adhesion. Conversely, a chimeric integrin construct combining the beta 8 extracellular domain with the beta 3 transmembrane and cytoplasmic domains did promote adhesion of transfected 293 cells. This suggests that the beta 8 cytoplasmic domain does not interact with the cytoskeleton and with cytoplasmic signaling pathways in an adhesion-promoting fashion. We conclude that the beta 8 cytoplasmic domain, which is structurally unrelated to the conserved cytoplasmic domains of other beta subunits, is functionally distinct.

Amino Acid Sequence↗
PubMed1994

Crystal structure of human angiogenin reveals the structural basis for its functional divergence from ribonuclease.

Angiogenin, a potent inducer of neovascularization, is the only angiogenic molecule known to exhibit ribonucleolytic activity. Its overall structure, as determined at 2.4 A, is similar to that of pancreatic ribonuclease A, but it differs markedly in several distinct areas, particularly the ribonucleolytic active center and the putative receptor binding site, both of which are critically involved in biological function. Most strikingly, the site that is spatially analogous to that for pyrimidine binding in ribonuclease A differs significantly in conformation and is "obstructed" by glutamine-117. Movement of this and adjacent residues may be required for substrate binding to angiogenin and, hence, constitute a key part of its mechanism of action.

Amino Acid Sequence↗
PubMed2005

Gene duplication and functional divergence during evolution of the cytoskeletal linker protein talin.

The animal talins are large, modular proteins that link the actin cytoskeleton to the extracellular environment through interactions with beta-integrins and actin. Dictyostelium discoideum has two talins, TalA and TalB, which have distinct physiological roles in cell adhesion, cell differentiation, and cytokinesis. We previously identified a second talin gene in vertebrates. Thus, talin function in vertebrates is also due to the action of multiple proteins. Using a phylogenomic approach we have determined that D. discoideum TalA/B and the animal talins are related by descent from a common ancestral talin and that duplication of TLN2 early in the chordate lineage produced TLN1. An additional duplication subsequently produced a second Talin-2 in teleost fishes and a second Talin-1 in Xenopus laevis. We also show that vertebrate Talin-2 mRNA is alternatively processed. In the invertebrate Drosophila melanogaster and in the non-vertebrate chordate Ciona intestinalis, which each have only one talin gene, alternative processing of talin mRNA also produces multiple talin species. Thus, in these organisms, talin function may be due to the action of more than one protein. To identify isoform-specific functions of vertebrate talins we have shown through proteomic analysis that mammalian Talin-1 and Talin-2 bind to different protein partners. Further characterization of the differences between animal talins, especially the direct comparison of talins in the model urochordate C. intestinalis, which has one talin gene that produces two talins through alternative mRNA splicing, with Talin-1 and Talin-2 in model vertebrates, will provide an experimental system for studying neofunctionalization or subfunctionalization of talin following the vertebrate talin gene duplication.

Amino Acid Sequence↗
PubMed1998

Structural and functional divergence of class II alcohol dehydrogenase--cloning and characterisation of rabbit liver isoforms of the enzyme.

cDNAs coding for class II alcohol dehydrogenase were isolated from a rabbit-liver cDNA library. Deduced amino acid sequences show that isozymic forms of rabbit class II alcohol dehydrogenase exist, with a positional identity of 88.4%. A high variability in structure of class II alcohol dehydrogenase between the species is also reflected in function. The rabbit II-1 isozyme shows common characteristics with the human enzyme, but has a lower Km value for ethanol, 4.2 mM. The II-2 isozyme shows restriction for aliphatic alcohols longer than pentanol. For shorter alcohols the II-2 form has similar Km values as the II-1 isozyme, 5.5 mM for ethanol, but is a low activity variant with a 10-fold decrease in k(cat) values compared with II-1. Nevertheless, II-2 has a higher specificity for benzoquinone than II-1 due to a lower Km value, 80 microM compared with 1 mM, and is in this sense more like the human class II enzyme. In addition a rabbit class III alcohol dehydrogenase cDNA was isolated that encodes a typical class III enzyme/glutathione-dependent formaldehyde dehydrogenase. The finding of isozymic forms of class II alcohol dehydrogenase is in line with the evolution of the system of medium-chain alcohol dehydrogenases with different enzymes, different classes and different isozymes and further underline the complexity of the entire mammalian alcohol dehydrogenase system.

Alcohol Dehydrogenase↗
PubMed1999

Functional divergence of the plastid and cytosolic forms of the 54-kDa subunit of signal recognition particle.

Chloroplast and cytoplasmic signal recognition particles (cpSRP and cySRP) each contain a similar subunit, SRP54. The chloroplast homologue binds to cpSRP43, which is absent from cytosolic SRP, and cySRP54 binds to SRP-RNA, which appears to be absent from cpSRP. In the presence of cpSRP43, cpSRP54 posttranslationally forms a soluble targeting intermediate, transit complex, with the major light harvesting protein of the thylakoid membrane. In contrast, cySRP54 functions cotranslationally. In this study we investigated whether cytosolic and chloroplast forms of SRP54 were interchangeable in three types of functional assays: complementation of an Escherichia coli SRP54 mutant, formation of the transit complex, and heterologous binding between the SRP54 subunits, cpSRP43, and SRP-RNA. In no cases were the 54-kDa subunits able to substitute for each other suggesting that the two proteins are fundamentally different.

Amino Acid Sequence↗
PubMed2006

The major human and mouse granzymes are structurally and functionally divergent.

Approximately 2% of mammalian genes encode proteases. Comparative genomics reveals that those involved in immunity and reproduction show the most interspecies diversity and evidence of positive selection during evolution. This is particularly true of granzymes, the cytotoxic proteases of natural killer cells and CD8+ T cells. There are 5 granzyme genes in humans and 10 in mice, and it is suggested that granzymes evolve to meet species-specific immune challenge through gene duplication and more subtle alterations to substrate specificity. We show that mouse and human granzyme B have distinct structural and functional characteristics. Specifically, mouse granzyme B is 30 times less cytotoxic than human granzyme B and does not require Bid for killing but regains cytotoxicity on engineering of its active site cleft. We also show that mouse granzyme A is considerably more cytotoxic than human granzyme A. These results demonstrate that even "orthologous" granzymes have species-specific functions, having evolved in distinct environments that pose different challenges.

Amino Acid Sequence↗
PubMed1996

Duplication and functional divergence in the chalcone synthase gene family of Asteraceae: evolution with substrate change and catalytic simplification.

Plant-specific polyketide synthase genes constitute a gene superfamily, including universal chalcone synthase [CHS; malonyl-CoA:4-coumaroyl-CoA malonyltransferase (cyclizing) (EC 2.3.1.74)] genes, sporadically distributed stilbene synthase (SS) genes, and atypical, as-yet-uncharacterized CHS-like genes. We have recently isolated from Gerbera hybrida (Asteraceae) an unusual CHS-like gene, GCHS2, which codes for an enzyme with structural and enzymatic properties as well as ontogenetic distribution distinct from both CHS and SS. Here, we show that the GCHS2-like function is encoded in the Gerbera genome by a family of at least three transcriptionally active genes. Conservation within the GCHS2 family was exploited with selective PCR to study the occurrence of GCHS2-like genes in other Asteraceae. Parsimony analysis of the amplified sequences together with CHS-like genes isolated from other taxa of angiosperm subclass Asteridae suggests that GCHS2 has evolved from CHS via a gene duplication event that occurred before the diversification of the Asteraceae. Enzyme activity analysis of proteins produced in vitro indicates that the GCHS2 reaction is a non-SS variant of the CHS reaction, with both different substrate specificity (to benzoyl-CoA) and a truncated catalytic profile. Together with the recent results of Durbin et al. [Durbin, M. L., Learn, G. H., Jr., Huttley, G. A. & Clegg, M. T. (1995) Proc. Natl. Acad. Sci. USA 92, 3338-3342], our study confirms a gene duplication-based model that explains how various related functions have arisen from CHS during plant evolution.

Acyl Coenzyme A↗
PubMed1998

CD46 (membrane cofactor protein of complement, measles virus receptor): structural and functional divergence among species (review).

Human CD46 was identified as a complement regulator and was later shown to be a measles virus receptor. The ubiquitous distribution profile of CD46 accounted for systemic measles infection and general protection of host tissue/organs from autologous complement. A similar ubiquitous distribution was observed for swine and simian CD46 homologues based upon subsequent cDNA cloning and Northern analysis, reinforcing the roles of CD46. In contrast, recent cDNA cloning and distribution analyses of murine and guinea-pig CD46 revealed the predominant expression of these rodent CD46 homologues in the testis, especially in mature testicular germ cells. These results do not support the established functions of human CD46 but support the hypothesis that CD46 on sperm serves as a fertilization-related adhesion molecule toward eggs. Here, we review the structure, function and distribution of human CD46 and discuss the possible differences between human CD46 and its homologues recently cloned from a variety of non-human primates and other animals.

Amino Acid Sequence↗
PubMed1999

Functional divergence between the half-sites of the DNA-binding sequence for the yeast transcriptional regulator Rap1p.

The yeast transcriptional regulator Rap1p binds to the DNA consensus sequence ACACCCAYACAYYY. We have previously shown that DNA-binding sites in which all four Y (Y=T or C) positions were Ts (UASrpg sequences) synergized more efficiently to activate transcription than sequences in which all Ys were Cs (telomere sequences) [F.-Z. Idrissi, J. Fernández-Larrea and B. Piña (1998) J. Mol. Biol. 284, 925-935]. Here we provide evidence that the DNA consensus sequence for Rap1p behaves as a combination of two ACAYYY half-sites with different functionality, the presence of Ts in the second half-site being the determinant for the transcriptional behaviour of the UASrpg sequences. DNA structure in the different complexes with Rap1p varied from being relatively uniform to appear rather distorted, this also being dependent on the presence of Ts in the second half-site. These distortions did not cause sharp bends or kinks in the DNA molecule. Computer analysis suggests that high-affinity binding of Rap1p to UASrpg sequences requires a rearrangement of the C-terminal Myb domain of the protein. We propose that the structural alterations in Rap1p-DNA complexes, both in the DNA and in the protein, affect the transcription potential of the complex in an allosteric manner. We also propose that the dimeric nature of the Rap1 DNA-binding domain is a key structural feature that explains the disparate functions of its DNA-binding sites in vivo.

Base Sequence↗
PubMed2005

Functional divergence of duplicated c-myc genes in a tetraploid fish, the common carp (Cyprinus carpio).

The proto-oncogene c-myc is thought to be one of the most important genes in controlling cell proliferation. In a tetraploid fish, two c-myc genes (CAM1 and CAM2) were previously isolated from the common carp, Cyprinus carpio, and were shown to have different expression patterns in adult tissues. Here we found that CAM1 and CAM2 proteins had distinct properties in terms of their transcription regulation system, formation of the transcription activator complex Myc/Max, and transcriptional activation of the target gene. These results showed that the two carp c-Myc proteins have overlapping but distinct functions, suggesting that CAM1 and CAM2 are evolving to acquire different functions after an earlier tetraploidization event.

Animals↗
PubMed1982

Mouse kallikrein arginyl-esteropeptidase genes: analysis of cloned cDNAs suggests rapid functional divergence from a common ancestral sequence.

A previously-cloned cDNA coding for a member of the kallikrein arginyl-esteropeptidase group of serine proteases, (pMK-1), was used as a hybridization probe to identify a second partial cDNA clone (pMK-2) from mouse submaxillary gland. pMK-2 shares more than a 98% nucleotide sequence homology with pMK-1; the 3' untranslated regions are identical and there are only two predicted amino acid changes over the C-terminal 66 amino acids. The site of one change is implicated in determining substrate specificity, while the other may affect the catalytic mechanism. Thus despite the marked similarity of pMK-1 and pMK-2, the differences probably give rise to functionally distinct enzymes and are not simply polymorphic alleles.

Amino Acid Sequence↗
PubMed2005

Functional divergence of Kaposi's sarcoma-associated herpesvirus and related gamma-2 herpesvirus thymidine kinases: novel cytoplasmic phosphoproteins that alter cellular morphology and disrupt adhesion.

The nucleoside kinase encoded by Kaposi's sarcoma-associated herpesvirus (KSHV) is a relatively inefficient enzyme with substrate specificity for thymidine alone, unlike alphaherpesvirus thymidine kinases (TKs). Similar to all gammaherpesvirus TKs, KSHV TK is composed of two distinct domains, a conserved C-terminal kinase and a novel and uncharacterized N terminus. Ectopic expression of KSHV TK in adherent cells induced striking morphological changes and anchorage independence although cells survived, a property shared with the related rhadinovirus TKs of rhesus monkey rhadinovirus and herpesvirus saimiri. To determine whether KSHV TK served alternate functions relevant to the rhadinovirus life cycle and to reveal the contribution of the N terminus, an enhanced green fluorescent protein-tagged fusion protein and serial mutants were generated for investigation of intracellular localization and cell biology. Analysis of truncation mutants showed that a proline-rich region located within the N terminus cooperated with the conserved C-terminal kinase to tether KSHV TK to a reticular network in the cytoplasm and to induce morphological change. Fusion of the KSHV N terminus to herpes simplex virus type 1 TK, a nucleus-localized enzyme, similarly resulted in cytoplasmic redistribution of the chimeric protein but did not alter cell shape or adhesion. Unlike other human herpesvirus TKs, KSHV TKs and related rhadinovirus TKs are constitutively tyrosine phosphorylated; a KSHV TK mutant that was hypophosphorylated failed to detach and grow in suspension. Loss of adhesion may enhance terminal differentiation, viral replication, and egress at the cellular level and at the organism level may facilitate detachment and distant migration of KSHV-replicating cells within body fluids--promoting oropharyngeal transmission and perhaps contributing to the multifocal lesions that characterize KS.

Cell Adhesion↗
PubMed2004

Functional divergence of antigen-specific T-lymphocyte responses in syngeneic graft-versus-host disease.

Administration of cyclosporine after autologous bone marrow transplantation elicits a T-lymphocyte autoaggression syndrome with remarkable similarity to graft-versus-host disease (GVHD). This syndrome, termed syngeneic GVHD (SGVHD), with both acute and chronic phases, is mediated by a restricted repertoire of autoreactive T cells that promiscuously recognize major histocompatibility complex (MHC) class II determinants in association with a peptide from the invariant chain (MHC class II-invariant chain peptide; CLIP). This study evaluated and compared antigen-specific autoreactive T cells during acute and chronic SGVHD ex vivo isolated with a soluble MHC class II immunoglobulin molecular construct. This approach allows for the direct assessment of the functional behavior of the effector T cells without potential modification by in vitro culture. Two major subsets were detected that had overlapping specificity recognizing the MHC class II-binding domain of CLIP but that were differentially dependent on the N- and C-terminal flanking domains of this peptide. The N- and C-terminal subsets were primarily associated with acute and chronic SGVHD, respectively. The cytokine profiles of the CLIP-reactive T cells, however, were most informative and closely correlated with the onset and progression of disease. Levels of type 1 cytokine, particularly interferon-gamma, messenger RNA (mRNA) production, assessed by quantitative polymerase chain reaction, were dominant during acute SGVHD, whereas chronic SGVHD was associated with type 2 cytokine mRNA production. Although there was a dramatic polarization with respect to cytokine production, only subtle changes in antigen specificity could be detected. Unexpectedly, the functional behavior within the antigen-specific effector cell populations is not fixed and seems to change as the disease progresses to the chronic phase. Concordant with the evolution of the effector T-cell response is a differential loss in B7.1 mRNA expression in the N-terminal CLIP-reactive T-cell subset that may reflect the regulation of this autoimmune response. Of additional interest, autoreactive T cells producing interleukin-10 mRNA were detected in both acute and chronic SGVHD, suggesting that this cytokine may play an important but perhaps paradoxical role in both the onset and progression of this experimental autoaggression syndrome.

Amino Acid Sequence↗
PubMed2000

Molecular cloning of an N-terminal splice variant of the capsaicin receptor. Loss of N-terminal domain suggests functional divergence among capsaicin receptor subtypes.

Recently a cDNA clone, vanilloid receptor subtype-1 (VR1), was isolated and found to encode an ion channel that is activated by both capsaicin, the pain producing compound in chili peppers, and by noxious thermal stimuli. Subsequently, two related cDNAs have been isolated, a stretch inactivating channel with mechanosensitive properties and a vanilloid receptor-like protein that is responsive to high temperatures (52-53 degrees C). Here, we report the isolation of a vanilloid receptor 5'-splice variant (VR.5'sv) which differs from VR1 by elimination of the majority of the intracellular N-terminal domain and ankyrin repeat elements. Both VR.5'sv and VR1 mRNA were shown to be expressed in tissues reportedly responsive to capsaicin including dorsal root ganglion, brain, and peripheral blood mononuclear cells. Functional expression of VR.5'sv in Xenopus oocytes and mammalian cells showed no sensitivity to capsaicin, the potent vanilloid resiniferatoxin, hydrogen ions (pH 6.2), or noxious thermal stimuli (50 degrees C). Since VR.5'sv is otherwise identical to VR1 throughout its transmembrane spanning domains and C-terminal region, these results support the hypothesis that the N-terminal intracellular domain is essential for the formation of functional receptors activated by vanilloid compounds and noxious thermal stimuli.

Amino Acid Sequence↗
PubMed2005

Different expression patterns of duplicated PHANTASTICA-like genes in Lotus japonicus suggest their divergent functions during compound leaf development.

Recent studies on leaf development demonstrate that the mechanism on the adaxial-abaxial polarity pattern formation could be well conserved among the far-related species, in which PHANTASTICA (PAHN)-like genes play important roles. In this study, we explored the conservation and diversity on functions of PHAN-like genes during the compound leaf development in Lotus japonicus, a papilionoid legume. Two PHAN-like genes in L. japonicus, LjPHANa and LjPHANb, were found to originate from a gene duplication event and displayed different expression patterns during compound leaf development. Two mutants, reduced leaflets1 (rel1) and reduced leaflets3 (rel3), which exhibited decreased adaxial identity of leaflets and reduced leaflet initiation, were identified and investigated. The expression patterns of both LjPHANs in rel mutants were altered and correlated with abnormalities of compound leaves. Our data suggest that LjPHANa and LjPHANb play important but divergent roles in regulating adaxial-abaxial polarity of compound leaves in L. japonicus.

Amino Acid Sequence↗
PubMed2000

Functional divergence of human genioglossus motor units with respiratory-related activity.

The genioglossus muscle has at least two types of motor unit with respiratory-related activity. Inspiratory motor units show phasic activity during inspiration, whereas inspiratory/expiratory motor units show phasic inspiratory activity superimposed on tonic activity. The purpose of this study was to investigate the physiological roles of these different genioglossus motor units. The unitary activities of 12 inspiratory and 12 inspiratory/expiratory motor units were recorded using fine-wire electrodes during quiet nasal breathing in eight normal adult males. The mean interspike interval and the SD of successive spikes were calculated for inspiratory and inspiratory/expiratory motor units, respectively. Scattergrams of the mean interspike interval versus SD were constructed for the two groups of motor units. The effects of changes in head position on the firing activity and the patterns of distribution of the mean interspike interval versus its SD were significantly different for inspiratory and inspiratory/expiratory motor units. These results suggest that the inspiratory and inspiratory/expiratory motor units have different functional roles in respiration; inspiratory motor units may be phasically active to counteract intraluminal negative pressure during inspiration, whereas inspiratory/expiratory motor units may be tonically active to maintain tongue posture.

Action Potentials↗
PubMed2000

Netrin-G1: a novel glycosyl phosphatidylinositol-linked mammalian netrin that is functionally divergent from classical netrins.

UNC-6/netrins compose a small phylogenetically conserved family of proteins that act as axon guidance cues. With a signal sequence trap method, we isolated a cDNA encoding a novel member of the UNC-6/netrin family, which we named netrin-G1. Unlike classical netrins, netrin-G1 consists of at least six isoforms of which five were predominantly anchored to the plasma membrane via glycosyl phosphatidyl-inositol linkages. Netrin-G1 transcripts were first detected in midbrain and hindbrain regions by embryonic day 12 and reached highest levels at perinatal stages in various brain regions, including olfactory bulb mitral cells, thalamus, and deep cerebellar nuclei. Its expression was primarily restricted to the CNS. Interestingly, netrin-G1 proteins did not show appreciable affinity to any netrin receptor examined. Unlike netrin-1, a secreted form of netrin-G1 consistently failed to attract circumferentially growing axons from the cerebellar plate. Our findings suggest that netrin-G1 and its putative receptors have coevolved independently from the classical netrins. The expression pattern of netrin-G1 and its predicted neuronal membrane localization suggest it may also have novel signaling functions in nervous system development.

Amino Acid Sequence↗
PubMed2003

Functional divergence of human cytoplasmic myosin II: kinetic characterization of the non-muscle IIA isoform.

Cytoplasmic (or non-muscle) myosin II isoforms are widely expressed molecular motors playing essential cellular roles in cytokinesis and cortical tension maintenance. Two of the three human non-muscle myosin II isoforms (IIA and IIB) have been investigated at the protein level. Transient kinetics of non-muscle myosin IIB showed that this motor has a very high actomyosin ADP affinity and slow ADP release. Here we report the kinetic characterization of the non-muscle myosin IIA isoform. Similar to non-muscle myosin IIB, non-muscle myosin IIA shows high ADP affinity and little enhancement of the ADP release rate by actin. The ADP release rate constant, however, is more than an order of magnitude higher than the steady-state ATPase rate. This implies that non-muscle myosin IIA spends only a small fraction of its ATPase cycle time in strongly actin-bound states, which is in contrast to non-muscle myosin IIB. Non-muscle myosin II isoforms thus appear to have distinct enzymatic properties that may be of importance in carrying out their cellular functions.

Actins↗