Actin Depolymerizing Factor Gene Research Articles

Genome-Wide Identification and Functional Validation of Actin Depolymerizing Factor (ADF) Gene Family in Gossypium hirsutum L.

The Actin Depolymerizing Factor (ADF) protein, highly conserved among eukaryotes, is essential for plant growth, development, and stress responses. Cotton, a vital economic crop with applications spanning oilseed, textiles, and military sectors, has seen a limited exploration of its ADF gene family. This research has identified 118 unique ADF sequences across four principal cotton species: Gossypium hirsutum L., Gossypium barbadense Linn, Gossypium raimondii, and Asiatic cotton. The study found that the structural domains and physicochemical properties of these proteins are largely uniform across species. The ADF genes were classified into four subfamilies with a notable expansion in groups III and IV due to tandem and chromosomal duplication events. A thorough analysis revealed a high degree of conservation in gene structure, including exon counts and the lengths of introns and exons, with the majority of genes containing three exons, aligning with the characteristics of the ADF family. RNA-seq analysis uncovered a spectrum of responses by GhADFs to various abiotic stresses with GhADF19 showing the most significant reaction. Virus-induced gene silencing (VIGS) experiments were conducted to assess the role of GhADF19 in plant growth under abiotic stress. The results demonstrated that plants with silenced GhADF19 exhibited significantly slower growth rates and lower dry weights when subjected to cold, salt, and drought stress compared to the control group. This marked reduction in growth and dry weight under stress conditions highlights the potential importance of GhADF19 in stress tolerance mechanisms.

The Role of the ADF Gene Family in Maize Response to Abiotic Stresses

The highly conserved actin depolymerizing factor (ADF) plays an important role in plant growth, development and responses to biotic and abiotic stresses. A total of 72 ADF genes in Arabidopsis, wheat, rice and sorghum can be divided into four groups. The multicollinearity analysis revealed that the maize ADF gene family exhibited more collinearity events with closely related gramineous plants. Fifteen ADF genes in maize were screened from the latest database, and bioinformatics analysis showed that these ADF genes were distributed across seven chromosomes in maize. The gene structure of the ADF gene family in maize exhibits significant conservation and cluster consistency. The promoter region contains rich regulatory elements that are involved in various regulations related to growth, development and adverse stresses. The drought-tolerant ZmADF5 gene in maize was further studied, and it was found that the allelic variations in ZmADF5 were mainly concentrated in its promoter region. A superior haplotype, with drought tolerance, was identified by candidate-gene association analysis of 115 inbred lines. By comparing the phenotypes of anthesis silking interval, grain yield and ear height, it was found that Hap2 performed better than Hap1 under drought stress. This study provides a theoretical reference for understanding the function of the ADF gene family and proposes further investigation into the role of ZmADF5 in abiotic-stress tolerance.

Overcoming functional redundancy in three actin depolymerizing factor genes for rice pollen tube growth.

Overcoming functional redundancy in three actin depolymerizing factor genes for rice pollen tube growth.

Actin-Depolymerizing Factor Gene Family Analysis Revealed That CsADF4 Increased the Sensitivity of Sweet Orange to Bacterial Pathogens.

The actin-depolymerizing factor (ADF) gene family regulates changes in actin. However, the entire ADF family in the sweet orange Citrus sinensis has not been systematically identified, and their expressions in different organs and biotic stress have not been determined. In this study, through phylogenetic analysis of the sweet orange ADF gene family, seven CsADFs were found to be highly conserved and sparsely distributed across the four chromosomes. Analysis of the cis-regulatory elements in the promoter region showed that the CsADF gene had the potential to impact the development of sweet oranges under biotic or abiotic stress. Quantitative fluorescence analysis was then performed. Seven CsADFs were differentially expressed against the invasion of Xcc and CLas pathogens. It is worth noting that the expression of CsADF4 was significantly up-regulated at 4 days post-infection. Subcellular localization results showed that CsADF4 was localized in both the nucleus and the cytoplasm. Overexpression of CsADF4 enhanced the sensitivity of sweet orange leaves to Xcc. These results suggest that CsADFs may regulate the interaction of C. sinensis and bacterial pathogens, providing a way to further explore the function and mechanisms of ADF in the sweet orange.

Genome-wide identification of actin-depolymerizing factor gene family and their expression patterns under various abiotic stresses in soybean (Glycine max).

The actin-depolymerizing factor (ADF) encoded by a family of genes is highly conserved among eukaryotes and plays critical roles in the various processes of plant growth, development, and stress responses via the remodeling of the architecture of the actin cytoskeleton. However, the ADF family and the encoded proteins in soybean (Glycine max) have not yet been systematically investigated. In this study, 18 GmADF genes (GmADF1 - GmADF18) were identified in the soybean genome and were mapped to 14 different chromosomes. Phylogenetic analysis classified them into four groups, which was confirmed by their structure and the distribution of conserved motifs in the encoded proteins. Additionally, 29 paralogous gene pairs were identified in the GmADF family, and analysis of their Ka/Ks ratios indicated their purity-based selection during the evolutionary expansion of the soybean genome. The analysis of the expression profiles based on the RNA-seq and qRT-PCR data indicated that GmADFs were diversely expressed in different organs and tissues, with most of them responding actively to drought- and salt-induced stresses, suggesting the critical roles played by them in various biological processes. Overall, our study shows that GmADF genes may play a crucial role in response to various abiotic stresses in soybean, and the highly inducible candidate genes could be used for further functional studies and molecular breeding in soybean.

Abiotic Stress-Induced Actin-Depolymerizing Factor 3 From Deschampsia antarctica Enhanced Cold Tolerance When Constitutively Expressed in Rice.

The Antarctic flowering plant Deschampsia antarctica is highly sensitive to climate change and has shown rapid population increases during regional warming of the Antarctic Peninsula. Several studies have examined the physiological and biochemical changes related to environmental stress tolerance that allow D. antarctica to colonize harsh Antarctic environments; however, the molecular mechanisms of its responses to environmental changes remain poorly understood. To elucidate the survival strategies of D. antarctica in Antarctic environments, we investigated the functions of actin depolymerizing factor (ADF) in this species. We identified eight ADF genes in the transcriptome that were clustered into five subgroups by phylogenetic analysis. DaADF3, which belongs to a monocot-specific clade together with cold-responsive ADF in wheat, showed significant transcriptional induction in response to dehydration and cold, as well as under Antarctic field conditions. Multiple drought and low-temperature responsive elements were identified as possible binding sites of C-repeat-binding factors in the promoter region of DaADF3, indicating a close relationship between DaADF3 transcription control and abiotic stress responses. To investigate the functions of DaADF3 related to abiotic stresses in vivo, we generated transgenic rice plants overexpressing DaADF3. These transgenic plants showed greater tolerance to low-temperature stress than the wild-type in terms of survival rate, leaf chlorophyll content, and electrolyte leakage, accompanied by changes in actin filament organization in the root tips. Together, our results imply that DaADF3 played an important role in the enhancement of cold tolerance in transgenic rice plants and in the adaptation of D. antarctica to its extreme environment.

Genome-Wide Identification and Low Temperature Responsive Pattern of Actin Depolymerizing Factor (ADF) Gene Family in Wheat (Triticum aestivum L.).

The actin depolymerizing factor (ADF) gene family, which is conserved in eukaryotes, is important for plant development, growth, and stress responses. Cold stress restricts wheat growth, development, and distribution. However, genome-wide identification and functional analysis of the ADF family in wheat is limited. Further, because of the promising role of ADF genes in cold response, there is need for an understanding of the function of this family on wheat under cold stress. In this study, 25 ADF genes (TaADFs) were identified in the wheat genome and they are distributed on 15 chromosomes. The TaADF gene structures, duplication events, encoded conversed motifs, and cis-acting elements were investigated. Expression profiles derived from RNA-seq data and real-time quantitative PCR analysis revealed the tissue- and temporal-specific TaADF expression patterns. In addition, the expression levels of TaADF13/16/17/18/20/21/22 were significantly affected by cold acclimation or freezing conditions. Overexpression of TaADF16 increased the freezing tolerance of transgenic Arabidopsis, possibly because of enhanced ROS scavenging and changes to the osmotic regulation in cells. The expression levels of seven cold-responsive genes were up-regulated in the transgenic Arabidopsis plants, regardless of whether the plants were exposed to low temperature. These findings provide fundamental information about the wheat ADF genes and may help to elucidate the regulatory effects of the encoded proteins on plant development and responses to low-temperature stress.

Genome-Wide Identification and Characterization of Actin-Depolymerizing Factor (ADF) Family Genes and Expression Analysis of Responses to Various Stresses in Zea Mays L.

Actin-depolymerizing factor (ADF) is a small class of actin-binding proteins that regulates the dynamics of actin in cells. Moreover, it is well known that the plant ADF family plays key roles in growth, development and defense-related functions. Results: Thirteen maize (Zea mays L., ZmADFs) ADF genes were identified using Hidden Markov Model. Phylogenetic analysis indicated that the 36 identified ADF genes in Physcomitrella patens, Arabidopsis thaliana, Oryza sativa japonica, and Zea mays were clustered into five groups. Four pairs of segmental genes were found in the maize ADF gene family. The tissue-specific expression of ZmADFs and OsADFs was analyzed using microarray data obtained from the Maize and Rice eFP Browsers. Five ZmADFs (ZmADF1/2/7/12/13) from group V exhibited specifically high expression in tassel, pollen, and anther. The expression patterns of 13 ZmADFs in seedlings under five abiotic stresses were analyzed using qRT-PCR, and we found that the ADFs mainly responded to heat, salt, drought, and ABA. Conclusions: In our study, we identified ADF genes in maize and analyzed the gene structure and phylogenetic relationships. The results of expression analysis demonstrated that the expression level of ADF genes was diverse in various tissues and different stimuli, including abiotic and phytohormone stresses, indicating their different roles in plant growth, development, and response to external stimulus. This report extends our knowledge to understand the function of ADF genes in maize.

Genome-Wide Identification, Characterization and Expression Profiling of ADF Family Genes in Solanum lycopersicum L.

The actin depolymerizing factor (ADF) proteins have growth, development, defense-related and growth regulatory functions in plants. The present study used genome-wide analysis to investigate ADF family genes in tomato. Eleven tomato ADF genes were identified and differential expression patterns were found in different organs. SlADF6 was preferentially expressed in roots, suggesting its function in root development. SlADF1, SlADF3 and SlADF10 were predominately expressed in the flowers compared to the other organs and specifically in the stamen compared to other flower parts, indicating their potential roles in pollen development. The comparatively higher expression of SlADF3 and SlADF11 at early fruit developmental stages might implicate them in determining final fruit size. SlADF5 and SlADF8 had relatively higher levels of expression five days after the breaker stage of fruit development, suggesting their possible role in fruit ripening. Notably, six genes were induced by cold and heat, seven by drought, five by NaCl, and four each by abscisic acid (ABA), jasmonic acid (JA) and wounding treatments. The differential expression patterns of the SlADF genes under different types of stresses suggested their function in stress tolerance in tomato plants. Our results will be helpful for the functional characterization of ADF genes during organ and fruit development of tomato under different stresses.

The Infection of Cucumber (Cucumis sativus L.) Roots by Meloidogyne incognita Alters the Expression of Actin-Depolymerizing Factor (ADF) Genes, Particularly in Association with Giant Cell Formation.

Cucumber (Cucumis sativus L.) is threatened by substantial yield losses due to the south root-knot nematode (Meloidogyne incognita). However, understanding of the molecular mechanisms underlying the process of nematode infection is still limited. In this study, we found that M. incognita infection affected the structure of cells in cucumber roots and treatment of the cytoskeleton inhibitor (cytochalasin D) reduced root-knot nematode (RKN) parasitism. It is known that Actin-Depolymerizing Factor (ADF) affects cell structure, as well as the organization of the cytoskeleton. To address the hypothesis that nematode-induced abnormal cell structures and cytoskeletal rearrangements might be mediated by the ADF genes, we identified and characterized eight cucumber ADF (CsADF) genes. Phylogenetic analysis showed that the cucumber ADF gene family is grouped into four ancient subclasses. Expression analysis revealed that CsADF1, CsADF2-1, CsADF2-2, CsADF2-3 (Subclass I), and CsADF6 (Subclass III) have higher transcript levels than CsADF7-1, CsADF7-2 (Subclass II genes), and CsADF5 (Subclass IV) in roots. Members of subclass I genes (CsADF1, CsADF2-1, CsADF2-2, and CsADF2-3), with the exception of CsADF2-1, exhibited a induction of expression in roots 14 days after their inoculation (DAI) with nematodes. However, the expression of subclass II genes (CsADF7-1 and CsADF7-2) showed no significant change after inoculation. The transcript levels of CsADF6 (Subclass III) showed a specific induction at 21 DAI, while CsADF5 (Subclass IV) was weakly expressed in roots, but was strongly up-regulated as early as 7 DAI. In addition, treatment of roots with cytochalasin D caused an approximately 2-fold down-regulation of the CsADF genes in the treated plants. These results suggest that CsADF gene mediated actin dynamics are associated with structural changes in roots as a consequence of M. incognita infection.

Characterization of the Pear (Pyrus ussuriensis) Actin-depolymerizing Factor PuADF During Fruit Ripening

The polymerization or depolymerization of the cytoskeleton is very important in various aspects of plant development. ADF (actin-depolymerizing factor) is a typical actin binding protein that can mediate the polymerization or depolymerization of the actin cytoskeleton, but its role in fruit ripening remains unclear. Here, we characterized an ADF gene, PuADF. The expression of PuADF was evaluated in young leaves, stems, flowers, and roots as well as in fruits. The expression of PuADF was induced by ethephon treatment but inhibited by 1-MCP treatment. To explore the network of PuADF function in ‘Nanguo’ pear fruits ripening, we screened a cDNA library from ‘Nanguo’ pear fruits using PuADF as bait. Two potential interactors of PuADF were identified, PuAS (Asparagine Synthetase) and PuDAD1 (Defender Against Death 1), both of which have been associated with PCD (Programmed Cell Death). This direct interaction was further confirmed by yeast two-hybrid analyses. The expression levels of PuAS and PuDAD1 were affected by ethylene. Our results indicated that PuADF is involved in ethylene-mediated fruit ripening and might be related to PCD during fruit ripening. The possible mechanisms are discussed.

TaADF7, an actin‐depolymerizing factor, contributes to wheat resistance against Puccinia striiformis f. sp. tritici

The actin cytoskeleton is involved in plant defense responses; however, the role of the actin-depolymerizing factor (ADF) family, which regulates actin cytoskeletal dynamics, in plant disease resistance, is largely unknown. Here, we characterized a wheat (Triticum aestivum) ADF gene, TaADF7, with three copies located on chromosomes1A, 1B, and 1D, respectively. All three copies encoded the same protein, although there were variations in 19 nucleotide positions in the open reading frame. Transcriptional expression of the three TaADF7 copies were all sharply elevated in response to avirulent Puccinia striiformis f.sp. tritici (Pst) infection, with similar expression patterns. TaADF7 regulated the actin cytoskeletal dynamics by targeting the actin cytoskeleton to execute actin binding/severing activities. When the TaADF7 copies were all silenced by virus-induced gene silencing, the growth of Pst hypha increased and sporadic urediniospores were observed, as compared with control plants, upon inoculation with avirulent Pst. In addition, the accumulation of reactive oxygen species (ROS) and the hypersensitive response (HR) were greatly weakened, whereas cytochalasinB partially rescued the HR in TaADF7 knock-down plants. Together, these findings suggest that TaADF7 is likely to contribute to wheat resistance against Pst infection by modulating the actin cytoskeletal dynamics to influence ROS accumulation and the HR.

Molecular characterization of an actin depolymerizing factor fromCryptocaryon irritans

Actin depolymerizing factors regulate actin dynamics involved in cellular processes such as morphogenesis, motility, development and infection. Here, a novel actin depolymerizing factor gene (CiADF 2 ) was cloned from the cDNA library of Cryptocaryon irritans, a parasitic ciliate causing cryptocaryonosis. The full-length cDNA of CiADF 2 was 531 bp. Its open reading frame (ORF) was 417 bp, encoding a polypeptide of 138 aa with typical features of the ADF/cofilin family. Reverse transcription-PCR suggested that CiADF 2 is expressed in all stages of the life cycle. After site-directed mutagenesis of a non-universal genetic code, the ORF was subcloned in Escherichia coli. The bacteria were induced with the addition of isopropylthio-β-D-galactoside to express a fusion protein of recombinant CiADF2 (rCiADF2) with glutathione S transferase. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot confirmed the predicted molecular mass of rCiADF2 of 16·2 kDa. A mouse antibody against rCiADF2 recognized native CiADF2, and rCiADF2 reacted with mouse antisera against C. irritans trophonts. CiADF2 was abundant in the plasma around cytostomes, suggesting that CiADF2 is involved in ciliate movement. Moreover, rCiADF2 showed F-actin binding and depolymerizing activity. This study will help to clarify the pathogenic biology of the parasite and develop effective control measures for cryptocaryonosis.

Comprehensive analysis of differentially expressed rice actin depolymerizing factor gene family and heterologous overexpression of OsADF3 confers Arabidopsis Thaliana drought tolerance.

BackgroundActin depolymerizing factors (ADFs) are small actin-binding proteins. Many higher-plant ADFs has been known to involve in plant growth, development and pathogen defense. However, in rice the temporal and spatial expression of OsADF gene family and their relationship with abiotic stresses tolerance is still unknown.ResultsHere we reported the first comprehensive gene expression profile analysis of OsADF gene family. The OsADF genes showed distinct and overlapping gene expression patterns at different growth stages, tissues and abiotic stresses. We also demonstrated that both OsADF1 and OsADF3 proteins were localized in the nucleus. OsADF1 and OsADF3 were preferentially expressed in vascular tissues. Under ABA or abiotic stress treatments, OsADF3::GUS activity was enhanced in lateral roots and root tips. Ectopically overexpressed OsADF3 conferred the mannitol- and drought-stress tolerance of transgenic Arabidopsis seedlings by increasing germination rate, primary root length and survival. Several drought-tolerance responsive genes (RD22, ABF4, DREB2A, RD29A, PIP1; 4 and PIP2; 6) were upregulated in transgenic Arabidopsis under drought stress.ConclusionsThese results suggested that OsADF gene family may participate in plant abiotic stresses response or tolerance and would facilitate functional validation of other OsADF genes.Electronic supplementary materialThe online version of this article (doi:10.1186/1939-8433-5-33) contains supplementary material, which is available to authorized users.

Immune response and protective efficacy against homologous challenge in BALB/c mice vaccinated with DNA vaccine encoding Toxoplasma gondii actin depolymerizing factor gene

A DNA vaccine (pVAX1-TgADF) encoding Toxoplasma gondii actin depolymerizing factor (ADF) gene was constructed and the immune response and protective efficacy of this vaccine against homologous challenge in BALB/c mice were evaluated. High titers of specific antibody and increases in the percentage of CD4 + and CD8 + T lymphocyte cells were observed from BALB/c mice vaccinated with pVAX1-TgADF ( P < 0.05), when PBS group was used as control. The survival time of BALB/c mice in pVAX1-TgADF group was longer than those in control groups. The numbers of brain cysts in the experimental BALB/c mice immunized with pVAX1-TgADF reduced significantly compared with those in PBS group ( P < 0.05), and the rate of reduction could reach to around 42.8%. These results suggested that the DNA vaccine pVAX1-TgADF could generate specific humoral and cellular immune responses, prolong survival times, and reduce brain cysts load against T. gondii infection in BALB/c mice.

Molecular cloning and characterization of an actin- depolymerizing factor gene in Hevea brasiliensis

Actin-depolymerizing factor (ADF) plays important roles in regulating actin dynamics by maintaining the optimum equilibrium between unpolymerized actin molecules and assembled actin filaments in different cellular processes. In this study, the first ADF gene in Hevea brasiliensis designated as HbADF, was isolated. The HbADF contained an open reading frame (ORF) encoding 139 amino acids. The deduced HbADF showed high identities to plants ADF proteins. Besides a conserved ADF domain, HbADF also contained putative actin and specific F-actin binding sites, phosphorylation site and possible CAM (calmodulin) combining region. The phylogenetic analysis indicated that HbADF was clustered in the subclass I. Being consistent with phylogenetic result, the expression of HbADF was constitutive. The HbADF transcripts were upregulated by ethephon and wounding treatments; whereas, HbADF was firstly induced, and then gradually downregulated by jasmonic acid. The expression profiles and characterizations of HbADF suggested that HbADF might be associated with latex regeneration and flow in H. brasiliensis.

Actin-depolymerizing factor of second-generation merozoite in Eimeria tenella: clone, prokaryotic expression, and diclazuril-induced mRNA expression

Actin depolymerizing factor (ADF) is an essential actin-binding protein that plays a key role in the control of actin dynamics and actin-based motility processes in intracellular parasites. To determine the effects of diclazuril on ADF gene of second-generation merozoites (mz-ADF) mRNA expression in Eimeria tenella, mz-ADF gene was cloned by RT-PCR from extracted RNA in second-generation merozoite of E. tenella and successfully expressed by pET-28a vector in Escherichia coli BL21(DE3). Results showed that the full length of the cloned cDNA sequence of the mz-ADF gene is 476 bp including an ORF of 375 bp. The sequence has 100% homology with a published sequence of sporozoite stage E. tenella ADF mRNA (GenBank EF195234.1). The recombinant protein was induced to be expressed by 1 mM isopropyl beta-D: -1-thiogalactopyranoside in vitro. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that 16.99 kDa fusion protein existed in solvable form. Compared with the infected/control group, mz-ADF mRNA expression level was downregulated by 63.86% in the infected/treatment group with the treatment of diclazuril. In conclusion, the data presented here indicate that mz-ADF gene participates in an important role in the invasion host of E. tenella. Downregulation of mz-ADF mRNA expression enrich the mechanism study of diclazuril on E. tenella.

Three cotton genes preferentially expressed in flower tissues encode actin-depolymerizing factors which are involved in F-actin dynamics in cells

To investigate whether the high expression levels of actin-depolymerizing factor genes are related to pollen development, three GhADF genes (cDNAs) were isolated and characterized in cotton. Among them, GhADF6 and GhADF8 were preferentially expressed in petals, whereas GhADF7 displayed the highest level of expression in anthers, revealing its anther specificity. The GhADF7 transcripts in anthers reached its peak value at flowering, suggesting that its expression is developmentally-regulated in anthers. The GhADF7 gene including the promoter region was isolated from the cotton genome. To demonstrate the specificity of the GhADF7 promoter, the 5′-flanking region, including the promoter and 5′-untranslated region, was fused with the GUS gene. Histochemical assays demonstrated that the GhADF7:GUS gene was specifically expressed in pollen grains. When pollen grains germinated, very strong GUS staining was detected in the elongating pollen tube. Furthermore, overexpression of GhADF7 gene in Arabidopsis thaliana reduced the viable pollen grains and, consequently, transgenic plants were partially male-sterile. Overexpression of GhADF7 in fission yeast (Schizosaccharomyces pombe) altered the balance of actin depolymerization and polymerization, leading to the defective cytokinesis and multinucleate formation in the cells. Given all the above results together, it is proposed that the GhADF7 gene may play an important role in pollen development and germination.

Actin-Depolymerizing Factor2-Mediated Actin Dynamics Are Essential for Root-Knot Nematode Infection ofArabidopsis

Reorganization of the actin and microtubule networks is known to occur in targeted vascular parenchymal root cells upon infection with the nematode Meloidogyne incognita. Here, we show that actin-depolymerizing factor (ADF) is upregulated in the giant feeding cells of Arabidopsis thaliana that develop upon nematode infection and that knockdown of a specific ADF isotype inhibits nematode proliferation. Analysis of the levels of transcript and the localization of seven ADF genes shows that five are upregulated in galls that result from the infection and that ADF2 expression is particularly increased between 14 and 21 d after nematode inoculation. Further analysis of ADF2 function in inducible RNA interference lines designed to knock down ADF2 expression reveals that this protein is required for normal cell growth and plant development. The net effect of decreased levels of ADF2 is F-actin stabilization in cells, resulting from decreased F-actin turnover. In nematode-infected plants with reduced levels of ADF2, the galls containing the giant feeding cells and growing nematodes do not develop due to the arrest in growth of the giant multinucleate feeding cells, which in turn is due to an aberrant actin network.

Roles of the first intron on the expression of Arabidopsis ( Arabidopsis thaliana) genes for actin and actin-binding proteins

We have recently reported that intron is required for strong and constitutive expression of two actin-binding proteins (ABPs), actin-depolymerizing factor (ADF) and profilin in vegetative tissues. To examine whether the expression of their target gene, actin, is also regulated by the introns, we examined the role of the first intron on the expression of the ACT2 gene, which encodes vegetative actin, using transgenic plants and a transient expression system. Transgenic plants carrying the ACT2 promoter-GUS with its first intron showed strong expression throughout the vegetative tissues, and the intron was necessary for expression in the shoot apical meristem region and root tips. Along with changes in expression pattern, the intron also increased GUS expression about 13-fold. However, this enhancement was not observed when the intron was moved upstream of the ACT2 promoter or the minimal promoter, suggesting that the intron is not a transcriptional enhancer. To further examine the role of the intron in the expression of actin and ABP genes, introns of actin, ADF, and profilin genes were exchanged. Introns of vegetative profilin and the ADF genes significantly enhanced gene expression under the control of heterologous promoters. Together with the role of the first intron of the ACT2 gene, these results suggest that the expression of actin and ABPs is regulated by intron-mediated enhancement.