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At least 19 recordsLinked to original sources

PubMed2003

Xkid is degraded in a D-box, KEN-box, and A-box-independent pathway.

During mitosis, the Xenopus chromokinesin Kid (Xkid) provides the polar ejection forces needed at metaphase for chromosome congression, and its degradation is required at anaphase to induce chromosome segregation. Despite the fact that the degradation of Xkid at anaphase seems to be a key regulatory factor to induce chromosome movement to the poles, little is known about the mechanisms controlling this proteolysis. We investigated here the degradation pathway of Xkid. We demonstrate that Xkid is degraded both in vitro and in vivo by APC/Cdc20 and APC/Cdh1. We show that, despite the presence of five putative D-box motifs in its sequence, Xkid is proteolyzed in a D-box-independent manner. We identify a domain within the C terminus of this chromokinesin, with sequence GxEN, whose mutation completely stabilizes this protein by both APC/Cdc20 and APC/Cdh1. Moreover, we show that this degradation sequence acts as a transposable motif and induces the proteolysis of a GST-GXEN fusion protein. Finally, we demonstrate that both a D-box and a GXEN-containing peptides completely block APC-dependent degradation of cyclin B and Xkid, indicating that the GXEN domain might mediate the recognition and association of Xkid with the APC.

Amino Acid Motifs↗

PubMed2003

Solution structure and DNA binding property of the fifth HMG box domain in comparison with the first HMG box domain in human upstream binding factor.

Human upstream binding factor (hUBF) is a nucleolar transcription factor involved in transcription by RNA polymerase I. It contains six HMG box domains. The contribution of each HMG box motif to its function is different. hUBF HMG box 1 shows a very strong binding affinity for both the four-way DNA junction and a 15 bp GC-rich rRNA gene core promoter fragment, but hUBF HMG box 5 shows a much weaker binding affinity for the four-way DNA junction and the GC-rich rRNA gene core promoter fragment. To illustrate the molecular basis of their DNA binding difference, the solution structure of box 5 was studied by NMR. The tertiary structure of box 5 shows a common flattened L-shaped fold, similar to box 1 and other HMG boxes with known structures. It is formed by intersection of three helical arms: helix 1 (residues 9-25) and helix 2 (residues 30-42) pack into each other to form the major wing, while helix 3 (residues 48-70) is aligned with the extended N-terminal segment to form the minor wing. A hydrophobic core is formed by three tryptophans (W14, W41, and W52) to maintain the fold. Although there is similarity between the two structures, negative charged electrostatic surface potential in the concave face of the molecule of box 5 exhibits great difference compared to that of box 1 and other HMG boxes with known structures. That surface is involved in DNA binding. Besides, in positions which are involved in intercalating into a DNA base pair, there are hydrophobic residues in box 1 and other HMG boxes but polar residues in box 5. These differences may contribute to the loss of the DNA binding ability of box 5.

Amino Acid Sequence↗

PubMed1998

A 69-base pair fragment derived from human transcobalamin II promoter is sufficient for high bidirectional activity in the absence of a TATA box and an initiator element in transfected cells. Role of an E box in transcriptional activity.

A 69-base pair (bp) (-581/-513) fragment derived from human transcobalamin II distal promoter constructed upstream of a chloramphenicol acetyltransferase reporter gene demonstrated high bidirectional promoter activity in transfected epithelial Caco-2 cells. DNase I footprinting, gel mobility shift, supershift, and mutagenesis studies with the 69-bp fragment demonstrated that a GC box (-568/-559) and an E box (-523/-528), which interacted with Sp1/Sp3 and USF1/USF2 (where USF is upstream stimulatory factor), respectively, were required for the full transcriptional activity of this fragment. Whereas mutations in the GC box reduced the promoter activity by 50%, mutations in the E box alone or in both the E box and GC box resulted in 90% loss of transcriptional activity. The essential role of the E box in the bidirectional promoter activity was further demonstrated by transient transfection in Caco-2, K-562, and HeLa cells using a 29-bp (-541/-513) fragment that contained only the E box. Based on these results we suggest that 1) the E box is essential for both the GC box-dependent and -independent promoter activity of the 69-bp fragment, 2) cooperative interactions between Sp1/Sp3 and USFs are required for the full activation of the 69-bp promoter activity, and 3) the single E box is able to mediate bidirectional transcription in transfected cells in the absence of an obvious TATA box or a known initiator element.

Base Sequence↗

PubMed2000

Developmental expression of Y-box protein 1 mRNA and alternatively spliced Y-box protein 3 mRNAs in spermatogenic cells in mice.

Y-box proteins bind DNA and RNA and are characterized by a cold shock domain and a carboxyl-terminus containing clusters of aromatic and basic residues that alternate with clusters of acidic residues. Y-box proteins 1 and 3 in mouse testis were cloned here by 3' rapid amplification of cDNA ends (RACE) using a degenerate primer. Northern blots and reverse transcription-polymerase chain reaction (RT-PCR) established that the levels of Y-box protein 1 and 3 mRNAs are regulated individually: (i) Y-box protein 1 mRNA is strongly expressed in kidney, whereas Y-box protein 3 mRNA is strongly expressed in heart and muscle; (ii) Y-box protein 1 and 3 mRNAs are weakly expressed in early prepubertal testis and strongly expressed in pachytene spermatocytes, round spermatids, and elongated spermatids; and (iii) prepubertal testes and meiotic and haploid spermatogenic cells express two alternatively spliced Y-box protein 3 mRNAs encoding isoforms with different carboxyl termini, whereas somatic tissues primarily express one form. Sucrose gradients reveal that approximately 27% of both Y-box protein 3 mRNAs are translationally active in adult testis. In conclusion, spermatogenic cells in mice express five isoforms of Y-box proteins including Y-box protein 1, and two isoforms each of Y-box proteins 2 and 3. This multiplicity is intriguing because Y-box proteins are thought to activate transcription and repress translation in spermatogenic cells.

Alternative Splicing↗

PubMed2002

Contribution of the Box 1 and Box 2 motifs of cytokine receptors to Jak1 association and activation.

Kinases of the Jak family (Jak1/2/3 and Tyk2) interact with the membrane proximal domain of different cytokine receptors and play a critical role in the activation of cytokine and growth factor signaling pathways. In this report we demonstrate that both the Box 1 and Box 2 motif collaborate in the association and activation of Jak1 by type I interferons. Mutational analysis of the beta chain of type I interferon receptor (IFNalphaRbetaL/IFNAR2) revealed that Box 1 plays a more significant role in activation than in the association with Jak1. On the contrary, the Box 2 motif contributes more to the association with Jak1 than to kinase activation. Additionally, the study of the Jak1 binding sites on the IL2 receptor beta (IL2Rbeta), IFNgammaRalpha/IFNGR1, and IL10Ralpha/IL10R1 chains suggests that cytokine receptors have two different kinds of interaction with Jak1. One form of interaction involves the Box 1 and the previously described Box 2 motif, which we now designate as Box 2A, characterized by the VEVI and LEVL sequences present in IFNalphaRbetaL/IFNAR2 and IL2Rbeta subunits, respectively. The second form of interaction requires a motif termed Box 2B, which is present in the IFNgammaRalpha/IFNGR1 (SILLPKS) and IL10Ralpha/IL10R1 (SVLLFKK) chains. Interestingly, Box 2B localizes close to the membrane region (8-10 amino acids from the membrane) similar to Box 1, whereas Box 2A is more distal (38-58 amino acids from the membrane).

Amino Acid Sequence↗

PubMed1998

Operation Red Box: a pilot project of needle and syringe drop boxes for injection drug users in East Baltimore.

We assessed the acceptability and the use of a community-based needle and syringe disposal project designed to serve injection drug users. In June 1996, three surplus U.S. mail collection boxes were painted red and used as syringe and needle drop boxes in locations with high drug use in East Baltimore. Acceptance of the drop boxes was measured by focus groups of residents, drug users, and police, held before and after project implementation. Use was measured by weekly counts of needles recovered from the red boxes. A sample of all deposited needles was randomly chosen for needle washing and subsequent HIV antibody testing. Community impact was measured by systematic surveys of needles discarded on public sidewalks, in areas with and areas without drop boxes. Before implementation, members of focus groups expressed concerns that drop boxes could convey mixed messages to youth (e.g., seeming to condone drug use), might result in increased loitering, and could further community stigmatization. After project implementation, all focus groups expressed support of project expansion. In the first 10 months, 2971 needles were collected. Of 156 needles tested, 10.9% were positive for HIV antibody. Needle counts on the street showed no significant change in red box areas compared with control areas. In this pilot project, red boxes were accepted by the community and drug users. Police officers also used the boxes to dispose of confiscated needles. Although limited in the number of drop boxes and follow-up time, this pilot project shows promise as a community-based method of safe needle disposal.

Adolescent↗

PubMed2001

D box and KEN box motifs in budding yeast Hsl1p are required for APC-mediated degradation and direct binding to Cdc20p and Cdh1p.

The precise order of molecular events during cell cycle progression depends upon ubiquitin-mediated proteolysis of cell cycle regulators. We demonstrated previously that Hsl1p, a protein kinase that inhibits the Swe1p protein kinase in a bud morphogenesis checkpoint, is targeted for ubiquitin-mediated turnover by the anaphase-promoting complex (APC). Here, we investigate regions of Hsl1p that are critical both for binding to the APC machinery and for APC-mediated degradation. We demonstrate that Hsl1p contains both a destruction box (D box) and a KEN box motif that are necessary for Hsl1p turnover with either APC(Cdc20) or APC(Cdh1). In coimmunoprecipitation studies, the D box of full-length Hsl1p was critical for association with Cdc20p, whereas the KEN box was important for association with Cdh1p. Fusion of a 206-amino-acid fragment of Hsl1p containing these motifs to a heterologous protein resulted in APC-dependent degradation of the fusion protein that required intact D box and KEN box motifs. Finally, this bacterially expressed Hsl1p fusion protein interacted with Cdc20p and Cdh1p either translated in vitro or expressed in and purified from insect cells. Binding to Cdc20p and Cdh1p was disrupted completely by a D box/KEN box double mutant. These results indicate that D box and KEN box motifs are important for direct binding to the APC machinery, leading to ubiquitination and subsequent protein degradation.

Amino Acid Motifs↗

PubMed1995

Isolation of two soybean G-box binding factors which interact with a G-box sequence of an auxin-responsive gene.

G-box binding factors (GBFs) constitute a family of plant DNA-binding proteins that bind to the G-box motif, a regulatory cis element present in many plant genes with a palindromic DNA motif of CACGTG. Previously TCCACGTGTC, a G-box motif, from an auxin responsive gene GmAux28 has been identified as a sequence-specific protein-binding site. Here the isolation of two soybean cDNA clones, referred to as SGBF-1 and SGBF-2, encoding proteins which bind to the G-box motif is reported. The primary structure of SGBF-1 and SGBF-2 predicts that these proteins contain a basic leucine zipper (bZIP) DNA-binding domain and an N-terminal proline-rich domain. A dramatic difference in the pattern of protein-DNA complex formation was observed when recombinant SGBF-1 and SGBF-2 proteins were analyzed by electrophoretic mobility shift assays (EMSAs). The SGBF-1 binding pattern obtained with the G-box probe resulted in three major retarded bands while the SGBF-2 formed a single complex. This shows that the characteristically diffuse banding pattern of plant nuclear proteins interacting with the G-box is also observed in a binding assay using only one recombinant GBF. EMSAs were performed with a few selected binding sequences to study the effect of flanking nucleotides to the hexanucleotide G-box core motif. The binding specificity of the SGBF proteins resembles that described for type A cauliflower nuclear G-box binding proteins which bind class I G-box elements [(G/T)(C/A)CACGTG(G/T)(A/C)]. Phylogenetic analysis of 13 GBF-like proteins from various plant species reveals that the SGBF-1 and SGBF-2 proteins belong to different lineages, suggesting that they may have distinct functions in activating transcription.

Amino Acid Sequence↗

PubMed1999

Box H and box ACA are nucleolar localization elements of U17 small nucleolar RNA.

The nucleolar localization elements (NoLEs) of U17 small nucleolar RNA (snoRNA), which is essential for rRNA processing and belongs to the box H/ACA snoRNA family, were analyzed by fluorescence microscopy. Injection of mutant U17 transcripts into Xenopus laevis oocyte nuclei revealed that deletion of stems 1, 2, and 4 of U17 snoRNA reduced but did not prevent nucleolar localization. The deletion of stem 3 had no adverse effect. Therefore, the hairpins of the hairpin-hinge-hairpin-tail structure formed by these stems are not absolutely critical for nucleolar localization of U17, nor are sequences within stems 1, 3, and 4, which may tether U17 to the rRNA precursor by base pairing. In contrast, box H and box ACA are major NoLEs; their combined substitution or deletion abolished nucleolar localization of U17 snoRNA. Mutation of just box H or just the box ACA region alone did not fully abolish the nucleolar localization of U17. This indicates that the NoLEs of the box H/ACA snoRNA family function differently from the bipartite NoLEs (conserved boxes C and D) of box C/D snoRNAs, where mutation of either box alone prevents nucleolar localization.

Animals↗

PubMed1992

Accumulation of U14 small nuclear RNA in Saccharomyces cerevisiae requires box C, box D, and a 5', 3' terminal stem.

U14 is one of several nucleolar small nuclear RNAs required for normal processing of rRNA. Functional mapping of U14 from Saccharomyces cerevisiae has yielded a number of mutants defective in U14 accumulation or function. In this study, we have further defined three structural elements required for U14 accumulation. The essential elements include the U14-conserved box C and box D sequences and a 5', 3' terminal stem. The box elements are coconserved among several nucleolar small nuclear RNAs and have been implicated in binding of the protein fibrillarin. New mutational results show that the first GA bases of the box C sequence UGAUGA are essential, and two vital bases in box D have also been identified. An intragenic suppressor of a lethal box C mutant has been isolated and shown to contain a new box C-like PyGAUG sequence two bases upstream of normal box C. The importance of the terminal stem was confirmed from new compensatory base changes and the finding that accumulation defects in the box elements can be complemented by extending the terminal stem. The results suggest that the observed defects in accumulation reflect U14 instability and that protein binding to one or more of these elements is required for metabolic stability.

Base Composition↗

PubMed1990

The plant G box promoter sequence activates transcription in Saccharomyces cerevisiae and is bound in vitro by a yeast activity similar to GBF, the plant G box binding factor.

G box and I box sequences of the Arabidopsis thaliana ribulose-bisphosphate-1,5-carboxylase small subunit (RBCS) promoter are required for expression mediated by the Arabidopsis rbcS-1A promoter in transgenic tobacco plants and are bound in vitro by factors from plant nuclear extracts termed GBF and GA-1, respectively. We show here that a -390 to -60 rbcS-1A promoter fragment containing the G box and two I boxes activates transcription from a truncated iso-1-cytochrome c (CYC1) gene promoter in Saccharomyces cerevisiae. Mutagenesis of either the rbcS-1A G box or both I box sequences eliminated the expression mediated by this fragment. When polymerized, I box oligonucleotides were also capable of enhancing expression from the truncated CYC1 promoter. Single-copy G box sequences from the Arabidopsis rbcS-1A, Arabidopsis Adh and tomato rbcS-3A promoters were more potent activators and were used in mobility shift assays to identify a DNA binding activity in yeast functionally similar to GBF. In methylation interference experiments, the binding specificity of the yeast protein was indistinguishable from that obtained with plant nuclear extracts.

Base Sequence↗

PubMed1994

An E box in the desmin promoter cooperates with the E box and MEF-2 sites of a distal enhancer to direct muscle-specific transcription.

The first 85 nt upstream of the transcription initiation site of the mouse desmin gene, which contain an E box (E1), the binding site of the helix-loop-helix myogenic regulators, are sufficient to confer low level muscle-specific expression. High levels of desmin expression are due to an enhancer, located between nucleotides -798 and -976, which contains an additional E box (E2) and a muscle-specific enhancer factor-2 (MEF-2) binding site. We have previously shown that both myoD and myogenin can bind to the proximal (E1) and distal (E2) boxes. Here we demonstrate that MEF-2C, a myocyte-restricted member of the MEF-2 family, can bind to the desmin MEF-2 site. Functional units for the enhancer activity required intact E2 and MEF-2 elements. The desmin enhancer can function relatively well with either the E2 box or the MEF-2 site and only mutation of both eliminates transcriptional enhancement; the presence of both of these elements is required for maximum enhancer activity. On the other hand, mutagenesis of just the proximal E1 box showed that this element is essential for desmin gene expression. Double mutations of E1 with E2 or MEF-2 sites suggested that, to achieve high levels of desmin gene expression, E1 serves most possibly as an intermediary for either E2 or MEF-2 enhancer elements to function. The location of the E1 site relative to the TATA box is crucial. Its activity is DNA turn- and distance-dependent. Furthermore, this box seems to be the main element for desmin transactivation by myoD and myogenin in 10T1/2 cells. Its inactivation diminishes the transactivation by these factors; MRF4 and Myf5, however, can still partially function, possibly by using the distal E2 box.

Animals↗

PubMed2000

Identification of a novel SCAN box-related protein that interacts with MZF1B. The leucine-rich SCAN box mediates hetero- and homoprotein associations.

The SCAN box or leucine-rich (LeR) domain is a conserved motif found within a subfamily of C(2)H(2) zinc finger proteins. The function of a SCAN box is unknown, but it is predicted to form alpha-helices that may be involved in protein-protein interactions. Myeloid zinc finger gene-1B (MZF1B) is an alternatively spliced human cDNA isoform of the zinc finger transcription factor, MZF1. MZF1 and MZF1B contain 13 C(2)H(2) zinc finger motifs, but only MZF1B contains an amino-terminal SCAN box. A bone marrow cDNA library was screened for proteins interacting with the MZF1B SCAN box domain and RAZ1 (SCAN-related protein associated with MZF1B) was identified. RAZ1 is a novel cDNA that encodes a SCAN-related domain and arginine-rich region but no zinc finger motifs. Co-immunoprecipitation assays demonstrate that the SCAN box domain of MZF1B is necessary for association with RAZ1. By yeast two-hybrid analysis, the carboxyl terminus of RAZ1 is sufficient for interaction with the MZF1B SCAN box. Furthermore, MZF1B and RAZ1 each self-associate in vitro via a SCAN box-dependent mechanism. These data provide evidence that the SCAN box is a protein interaction domain that mediates both hetero- and homoprotein associations.

Alternative Splicing↗

PubMed1998

The role of the destruction box and its neighbouring lysine residues in cyclin B for anaphase ubiquitin-dependent proteolysis in fission yeast: defining the D-box receptor.

Programmed proteolysis of proteins such as mitotic cyclins and Cut2/Pds1p requires a 9-residue conserved motif known as the destruction box (D-box). Strong expression of protein fragments containing destruction boxes, such as the first 70 residues of Cdc13 (N70), inhibits the growth of Schizosaccharomyces pombe at metaphase. This inhibition can be overcome either by removal of all lysine residues from N70 using site-directed mutagenesis (K0-N70) or by raising the concentration of intracellular ubiquitin. Consistent with the idea that competition for ubiquitin accounts for some of its inhibitory effects, wild-type N70 not only stabilized D-box proteins, but also Rum1 and Cdc18, which are degraded by a different pathway. The K0-N70 construct was neither polyubiquitinated nor degraded in vitro, but it blocked the growth of strains of yeast in which anaphase-promoting complex/cyclosome (APC/C) function was compromised by mutation, and specifically inhibited proteolysis of APC/C substrates in vivo. Both K0-N70 and 20-residue D-box peptides blocked polyubiquitination of other D-box-containing substrates in a cell-free ubiquitination assay system. These data suggest the existence of a D-box receptor protein that recognizes D-boxes prior to ubiquitination.

Amino Acid Sequence↗

PubMed1991

C/EBP-like proteins binding to the functional box-alpha and box-beta of the second enhancer of hepatitis B virus.

The second enhancer (enhancer II) of hepatitis B virus is functionally liver specific. Located within an open reading frame of the virus and immediately upstream of the initiation sites of viral major transcripts, enhancer II furnishes a unique model for use in investigating the structure and function of an enhancer. In this study, two functional constituents, a 23-bp box-alpha and a 12-bp box-beta, are identified as being both necessary and sufficient for enhancer II function. Examination of the box-alpha and box-beta sequences reveals a weak homology to the extended consensus for a C/EBP binding site. Gel shift and footprinting analyses indicate that multiple proteins bind to these sequences and thus are candidate transcription factors that mediate the enhancer function. One heat-resistant protein, protein a, and one heat-sensitive protein, protein b, bind to box-alpha. Protein a, which binds to box-alpha in a way indistinguishable from that seen with a recombinant C/EBP, appears not to be identical to C/EBP in that the binding of protein a requires a minimal sequence larger than the canonical C/EBP sites. Two box-beta-binding proteins, c and d, show greater affinity for the C/EBP consensus than for box-beta. However, both proteins c and d are relatively heat sensitive and display a distinct sequence preference from the recombinant C/EBP protein. Since the function of enhancer II is strictly dependent on a bipartite architecture, this system provides a unique model for studies of how the interactions of its binding proteins lead to the enhancer function.

Base Sequence↗

PubMed2004

TBX5, a gene mutated in Holt-Oram syndrome, is regulated through a GC box and T-box binding elements (TBEs).

TBX5 is a member of the T-box gene family and encodes a transcription factor that regulates the expression of other gene(s) in the developing heart and limbs. Mutations of TBX5 cause Holt-Oram syndrome (HOS), an autosomal dominant condition characterized by congenital heart defects and limb anomalies. How TBX5 gene expression is regulated is still largely unknown. In order to identify transcription factors regulating TBX5 expression, we examined the 5'-flanking region of the human TBX5 gene. We determined that up to 300 bp of the 5'-flanking region of the TBX5 gene was necessary for promoter activity in mouse cardiomyocyte ECL2 cells. One GC box, three potential T-box-like binding elements (TBE-A, -B, and -C), and one NKX2.5 binding site were identified. Site-directed mutagenesis of the potential binding sites revealed that the GC box, TBE-B, TBE-C, and NKX2.5 are functionally positive for the expression of TBX5. DNA footprint analysis showed that these binding regions are resistant to DNaseI digestion. Electrophoretic mobility shift assays (EMSAs) further demonstrated the protein-DNA interactions at the GC box and the potential TBE-B, TBE-C, and NKX2.5 sites in a sequence-specific manner. The ability of TBX5 to regulate its own promoter was demonstrated by the ability of ectopically expressed human TBX5 to increase reporter expression. We conclude that the GC box, T-box-like binding elements, and NKX2.5 binding site play important roles in the regulation of TBX5 expression, and that TBX5 is likely to be autoregulated as part of the mechanism of its transcription.

5' Flanking Region↗

PubMed1990

Mutation of either G box or I box sequences profoundly affects expression from the Arabidopsis rbcS-1A promoter.

A deletion analysis of the Arabidopsis thaliana rbcS-1A promoter defined a 196 bp region (-320 to -125) sufficient to confer light-regulated expression on a heterologous Arabidopsis alcohol dehydrogenase (Adh) reporter gene in transgenic Nicotiana tabacum (tobacco) leaves. This region, which contains DNA sequences I, G and GT boxes, with homology to other ribulose-1,5-bisphosphate carboxylase small subunit (RBCS) gene promoter sequences, directed expression independent of orientation and relative position in the Adh promoter. Site-specific mutagenesis of these conserved sequences and subsequent expression analysis in transgenic tobacco showed that both G box and I box mutations in the context of the full (-1700 to +21) rbcS-1A promoter substantially reduced the expression of Adh and beta-glucuronidase (GUS) reporter genes. The G box has previously been shown to specifically bind in vitro a factor isolated from nuclear extracts of tomato and Arabidopsis. This factor (GBF) is distinct from the factor GT-1 which binds to adjacent GT boxes in the pea rbcS-3A promoter. Multiple mutations in putative Arabidopsis rbcS-1A promoter GT boxes had no pronounced affect on expression, possibly due to a redundancy of these sites. Experiments in which rbcS-1A promoter fragments were fused to truncated 35S CaMV (cauliflower mosaic virus) promoter--GUS reporter constructs showed that cis-acting CaMV promoter elements could partially restore expression to G-box-mutated rbcS-1A sequences.

Alcohol Dehydrogenase↗

PubMed1996

Classification and phylogeny of the MADS-box multigene family suggest defined roles of MADS-box gene subfamilies in the morphological evolution of eukaryotes.

The MADS-box encodes a novel type of DNA-binding domain found so far in a diverse group of transcription factors from yeast, animals, and seed plants. Here, our first aim was to evaluate the primary structure of the MADS-box. Compilation of the 107 currently available MADS-domain sequences resulted in a signature which can strictly discriminate between genes possessing or lacking a MADS-domain and allowed a classification of MADS-domain proteins into several distinct subfamilies. A comprehensive phylogenetic analysis of known eukaryotic MADS-box genes, which is the first comprising animal as well as fungal and plant homologs, showed that the vast majority of subfamily members appear on distinct subtrees of phylogenetic trees, suggesting that subfamilies represent monophyletic gene clades and providing the proposed classification scheme with a sound evolutionary basis. A reconstruction of the history of the MADS-box gene subfamilies based on the taxonomic distribution of contemporary subfamily members revealed that each subfamily comprises highly conserved putative orthologs and recent paralogs. Some subfamilies must be very old (1,000 MY or more), while others are more recent. In general, subfamily members tend to share highly similar sequences, expression patterns, and related functions. The defined species distribution, specific function, and strong evolutionary conservation of the members of most subfamilies suggest that the establishment of different subfamilies was followed by rapid fixation and was thus highly advantageous during eukaryotic evolution. These gene subfamilies may have been essential prerequisites for the establishment of several complex eukaryotic body structures, such as muscles in animals and certain reproductive structures in higher plants, and of some signal transduction pathways. Phylogenetic trees indicate that after establishment of different subfamilies, additional gene duplications led to a further increase in the number of MADS-box genes. However, several molecular mechanisms of MADS-box gene diversification were used to a quite different extent during animal and plant evolution. Known plant MADS-domain sequences diverged much faster than those of animals, and gene duplication and sequence diversification were extensively used for the creation of new genes during plant evolution, resulting in a relatively large number of interacting genes. In contrast, the available data on animal genes suggest that increase in gene number was only moderate in the lineage leading to mammals, but in the case of MEF2-like gene products, heterodimerization between different splice variants may have increased the combinatorial possibilities of interactions considerably. These observations demonstrate that in metazoan and plant evolution, increased combinatorial possibilities of MADS-box gene product interactions correlated with the evolution of increasingly complex body plans.

Amino Acid Sequence↗