Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “transcript”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 793 records · Page 44Linked to original sources

Transcriptional termination between bovine papillomavirus type 1 (BPV-1) early and late polyadenylation sites blocks late transcription in BPV-1-transformed cells.

Bovine papillomavirus type 1 (BPV-1) is a small DNA tumor virus which induces fibropapillomas in cattle and transforms rodent cells in culture. Transcripts are derived from a single strand of the circular viral genome, which has multiple promoters and two polyadenylation sites. In the transformed cell, the first (early) polyadenylation site is utilized exclusively and, therefore, only the early region is expressed. Transcription of the late genes, which requires use of the second (late) polyadenylation site, is seen only in the fully differentiated keratinocytes of the fibropapilloma. In this study, nascent RNA chain analysis of BPV-1-transformed C127 cells was used to demonstrate that at least 90% of the RNA polymerases which transcribe past the early polyadenylation site terminate transcription within the late region before reaching the late polyadenylation site. Therefore, transcription termination is at least partially responsible for the absence of late transcription in the BPV-1-transformed cell and is likely to be an important mechanism for regulation of papillomavirus late transcription during keratinocyte differentiation.

Animals↗

Investigation of the control of coronavirus subgenomic mRNA transcription by using T7-generated negative-sense RNA transcripts.

The subgenomic mRNAs of the coronavirus transmissible gastroenteritis virus (TGEV) are not produced in equimolar amounts. We have developed a reporter gene system to investigate the control of this differential subgenomic mRNA synthesis. Transcription of mRNAs by the TGEV polymerase was obtained from negative-sense RNA templates generated in situ from DNA containing a T7 promoter. A series of gene cassettes was produced; these cassettes comprised the reporter chloramphenicol acetyltransferase (CAT) gene downstream of transcription-associated sequences (TASs) (also referred to as intergenic sequences and promoters) believed to be involved in the synthesis of TGEV subgenomic mRNAs 6 and 7. The gene cassettes were designed so that negative-sense RNA copies of the CAT gene with sequences complementary to the TGEV TASs, or modified versions, at the 3' end would be synthesized in situ by T7 RNA polymerase. Using this system, we have demonstrated that CAT was expressed from mRNAs derived from the T7-generated negative-sense RNA transcripts only in TGEV-infected cells and only from transcripts possessing a TGEV negative-sense TAS. Analysis of the CAT mRNAs showed the presence of the TGEV leader RNA sequence at the 5' end, in keeping with observations that all coronavirus mRNAs have a 5' leader sequence corresponding to the 5' end of the genomic RNA. Our results indicated that the CAT mRNAs were transcribed from the in situ-synthesized negative-sense RNA templates without the requirement of TGEV genomic 5' or 3' sequences on the T7-generated negative-sense transcripts (3'-TAS-CAT-5'). Modification of the TGEV TASs indicated (i) that the degree of potential base pairing between the 3' end of the leader RNA and the TGEV negative-sense TAS was not the sole determinant of the amount of subgenomic mRNA transcribed and (ii) that other factors, including nucleotides flanking the TAS, are involved in the regulation of transcription of TGEV subgenomic mRNAs.

Animals↗

Human immunodeficiency virus type 1 integrase protein promotes reverse transcription through specific interactions with the nucleoprotein reverse transcription complex.

The human immunodeficiency virus type 1 (HIV-1) integrase protein (IN) is essential for integration of the viral DNA into host cell chromosomes. Since IN is expressed and assembled into virions as part of the 160-kDa Gag-Pol precursor polyprotein and catalyzes integration of the provirus in infected cells as a mature 32-kDa protein, mutations in IN are pleiotropic and may affect virus replication at different stages of the virus life cycle in addition to integration. Several different phenotypes have been observed for IN mutant viruses, including defects in virion morphology, protein composition, reverse transcription, nuclear import, and integration. Because the effects of mutations in the IN domain of Gag-Pol can not always be distinguished from those of mutations in the mature IN protein, there remains a significant gap in our understanding of IN function in vivo. To directly analyze the function of the mature IN protein itself, in the context of a replicating virus but independently from that of Gag-Pol, we used an approach developed in our laboratory for incorporating proteins into HIV virions by their expression in trans as fusion partners of either Vpr or Vpx. By providing IN in trans as a Vpr-IN fusion protein, our analysis revealed, for the first time, that the mature IN protein is essential for the efficient initiation of reverse transcription in infected cells and that this function does not require the IN protein to be enzymatically (integration) active. Our findings of a direct physical interaction between IN and reverse transcriptase and the failure of heterologous HIV-2 IN protein to efficiently support reverse transcription indicate that this novel function occurs through specific interactions with other viral components of the reverse transcription initiation complex. Studies involving complementation between integration- and DNA synthesis-defective IN mutants further support this conclusion and reveal that the highly conserved HHCC motif of IN is important for both activities. These findings provide important new insights into IN function and reverse transcription in the context of the nucleoprotein reverse transcription complex within the infected cell. Moreover, they validate a novel approach that obviates the need to mutate Gag-Pol in order to study the role of its individual mature components at the virus replication level.

DNA, Viral↗

The herpes simplex virus type 1 latency-associated transcript (LAT) enhancer/rcr is hyperacetylated during latency independently of LAT transcription.

During herpes simplex virus type 1 (HSV-1) latency, only one region of the viral genome is actively transcribed: the region encoding the latency-associated transcript (LAT). A previous study demonstrated that during latency the LAT promoter is hyperacetylated at histone H3 (K9, K14) relative to lytic genes examined. In the present study, we examine the acetylation profile of regions downstream of the LAT promoter during a latent infection of murine dorsal root ganglia. These analyses revealed the following: (i) the region of the genome containing the 5' exon of the LAT primary transcript was at least as enriched in acetylated H3 as the LAT promoter, and (ii) the region of hyperacetylation does not extend to the ICP0 promoter. In order to assess the contribution of LAT transcription to the acetylation of the 5' exon region, the acetylation profile of KOS/29, a recombinant with a deletion of the LAT promoter, was examined. The region containing the 5' exon of KOS/29 was hyperacetylated relative to lytic gene regions in the absence of detectable LAT transcription. These results indicate that the region containing the 5' exon of LAT, known to contain enhancer activities and to be critical for induced reactivation (rcr), exists in a chromatin structure during latency that is distinct from other lytic gene regions. This result suggests a role for the 5' exon LAT enhancer region as a cis-acting regulator of transcription that maintains a transcriptionally permissive chromatin domain in the HSV-1 latent episome.

Acetylation↗

Shutoff of RNA polymerase II transcription by poliovirus involves 3C protease-mediated cleavage of the TATA-binding protein at an alternative site: incomplete shutoff of transcription interferes with efficient viral replication.

The TATA-binding protein (TBP) plays a crucial role in cellular transcription catalyzed by all three DNA-dependent RNA polymerases. Previous studies have shown that TBP is targeted by the poliovirus (PV)-encoded protease 3C(pro) to bring about shutoff of cellular RNA polymerase II-mediated transcription in PV-infected cells. The processing of the majority of viral precursor proteins by 3C(pro) involves cleavages at glutamine-glycine (Q-G) sites. We present evidence that suggests that the transcriptional inactivation of TBP by 3C(pro) involves cleavage at the glutamine 104-serine 105 (Q104-S105) site of TBP and not at the Q18-G19 site as previously thought. The TBP Q104-S105 cleavage by 3C(pro) is greatly influenced by the presence of an aliphatic amino acid at the P4 position, a hallmark of 3C(pro)-mediated proteolysis. To examine the importance of host cell transcription shutoff in the PV life cycle, stable HeLa cell lines were created that express recombinant TBP resistant to cleavage by the viral proteases, called GG rTBP. Transcription shutoff was significantly impaired and delayed in GG rTBP cells upon infection with poliovirus compared with the cells that express wild-type recombinant TBP (wt rTBP). Infection of GG rTBP cells with poliovirus resulted in small plaques, significantly reduced viral RNA synthesis, and lower viral yields compared to the wt rTBP cell line. These results suggest that a defect in transcription shutoff can lead to inefficient replication of poliovirus in cultured cells.

3C Viral Proteases↗

Transcriptional analysis of latent and inducible Kaposi's sarcoma-associated herpesvirus transcripts in the K4 to K7 region.

Kaposi's sarcoma-associated herpesvirus (KSHV) is a gamma-2 herpesvirus with a genome containing a long unique coding region (LUR) flanked by GC-rich terminal repeat sequences. The LUR encodes approximately 90 annotated open reading frames (ORFs) with complex patterns of gene expression during viral latency, reactivation, and de novo infection. To identify unannotated KSHV genes, we examined the region between 21,500 and 30,000 bp of the KSHV LUR, representing approximately 8.5 kb of sequence. This region encodes seven known single-exon ORFs (K4, K4.1, K4.2, K5, K6, K7, and PAN), but previous computer analyses have failed to identify additional likely genes in the remaining 5.2 kb. We identified four novel transcripts using Northern blotting, phage library screening, and 5' rapid amplification of cDNA ends analysis in the region between ORFs K4.2 and K7. In vitro analysis of KSHV-infected primary effusion lymphoma cell lines in the presence of 12-O-tetradecanoylphorbol-13-acetate and phosphonoformic acid suggests that one latent transcript is coterminal with the previously annotated K3 gene encoding an ubiquitin-ligase known to downregulate major histocompatibility complex class I expression. This alternatively spliced transcript may contribute to KSHV adaptive immune evasion during latent infection. Other transcripts are inducible, including a 6.1-kb transcript that is the largest transcript found in the KSHV genome to date.

Cell Line↗

An in vitro transcription analysis of early responses of the human immunodeficiency virus type 1 long terminal repeat to different transcriptional activators.

In this report we introduce a simple, fast, and reliable method to prepare whole cell or nuclear extracts from small numbers of cells. These extracts were used to study transcriptional activation of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) in vitro. Our results revealed that the time courses of activation of extracts derived from cells stimulated with the mitogenic lectin phytohemagglutinin (PHA) or with the tumor promoter phorbol 12-myristate 13-acetate (PMA) are different. PMA induces a rapid onset of increased in vitro transcription from the HIV-1 LTR, while PHA causes a slow and sustained response. The biochemical relevance of protein synthesis inhibition by cycloheximide treatment of cells was investigated. In these studies, PMA induction of a change in in vitro transcriptional activity is not dependent on protein synthesis. Cycloheximide alone is insufficient to induce activation. Oligonucleotide-mediated site-directed mutagenesis demonstrated that mutation of the TATA box in the LTR ablated initiation of both basal-level transcription and activation by extracts from cells stimulated with PMA. Surprisingly, mutation of both kappa B sites in the LTR reduced but did not eliminate the in vitro response to extracts prepared at early time points after PHA or PMA stimulation of Jurkat cells. The reduction was greater in extracts derived from cells treated with PMA. Deletion analysis of the HIV-1 LTR revealed at least one region (-464 to -252) capable of suppressing in vitro transcription in extracts from Jurkat cells stimulated by PMA. This result is consistent with early studies of the HIV-1 LTR in transient transfection assays. We therefore have been able to observe distinct regulatory events at early time points after cells are exposed to agents known to induce transcription of both the HIV-1 LTR reporter gene constructs and the HIV-1 provirus itself.

Cell Line↗

Characterization of the inducer of short transcripts, a human immunodeficiency virus type 1 transcriptional element that activates the synthesis of short RNAs.

The inducer of short transcripts, or IST, is an unusual transcriptional element located downstream of the human immunodeficiency virus type 1 (HIV-1) promoter. IST activates HIV-1 transcription, but the resulting RNAs are short and end at approximately position +59. IST, therefore, appears to promote the formation of transcription complexes that are unable to elongate efficiently. This activity contrasts with that of TAR, the target for Tat trans-activation, which upon binding of the viral protein Tat promotes the formation of transcription complexes capable of efficient elongation through the entire viral genome. We have localized and characterized the IST element. Our results indicate that IST is located mainly between positions -5 and +26, although the sequences from positions +40 to +59 also contribute to IST activity. Unlike TAR, which is an RNA element, IST appears to be a DNA element. Thus, the HIV-1 R region is a complex regulatory region with RNA and DNA elements that promote the formation of transcription complexes with different elongation properties.

Base Sequence↗

Reactivation of apolipoprotein II gene transcription by cycloheximide reveals two steps in the deactivation of estrogen receptor-mediated transcription.

In this report, we describe apolipoprotein II (apoII) gene expression in cell lines derived by stable expression of the chicken estrogen receptor in LMH chicken hepatoma cells. In cell lines expressing high levels of receptor (LMH/2A), apoII gene expression is increased by estrogen 300-fold compared with levels in the receptor-deficient parent LMH line. LMH/2A cells show apoII mRNA induction and turnover kinetics similar to those in chicken liver. Inhibition of protein synthesis with cycloheximide (CHX) or puromycin following estrogen withdrawal superinduces apoII mRNA without affecting apoII mRNA stability. Superinduction is due to an estrogen-independent reactivation of apoII gene transcription. The apoII gene can be reactivated by CHX for up to 24 h following hormone withdrawal, suggesting that the gene is in a repressed yet transcriptionally competent state. These results reveal two distinct events necessary for termination of estrogen receptor-mediated transcription. The first event, removal of hormone, is sufficient to stop transcription when translation is ongoing. The second event is revealed by the CHX-induced superinduction of apoII mRNA following hormone withdrawal. This superinduction suggests that deactivation of estrogen receptor-mediated transcription requires a labile protein. Furthermore, reactivation of apoII gene expression by CHX and estrogen is additive, suggesting that estrogen is unable to overcome repression completely. Thus, a labile protein may act to repress estrogen receptor-mediated transcription of the apoII gene.

Animals↗

Implication of localization of human DNA repair enzyme O6-methylguanine-DNA methyltransferase at active transcription sites in transcription-repair coupling of the mutagenic O6-methylguanine lesion.

DNA lesions that halt RNA polymerase during transcription are preferentially repaired by the nucleotide excision repair pathway. This transcription-coupled repair is initiated by the arrested RNA polymerase at the DNA lesion. However, the mutagenic O6-methylguanine (6MG) lesion which is bypassed by RNA polymerase is also preferentially repaired at the transcriptionally active DNA. We report here a plausible explanation for this observation: the human 6MG repair enzyme O6-methylguanine-DNA methyltransferase (MGMT) is present as speckles concentrated at active transcription sites (as revealed by polyclonal antibodies specific for its N and C termini). Upon treatment of cells with low dosages of N-methylnitrosourea, which produces 6MG lesions in the DNA, these speckles rapidly disappear, accompanied by the formation of active-site methylated MGMT (the repair product of 6MG by MGMT). The ability of MGMT to target itself to active transcription sites, thus providing an effective means of repairing 6MG lesions, possibly at transcriptionally active DNA, indicates its crucial role in human cancer and chemotherapy by alkylating agents.

Alkylating Agents↗

A link between breast cancer and local estrogen biosynthesis suggested by quantification of breast adipose tissue aromatase cytochrome P450 transcripts using competitive polymerase chain reaction after reverse transcription.

C19 steroids are converted to estrogens in a number of tissues by a specific form of cytochrome P450, namely aromatase cytochrome P450 (P450arom). Adipose tissue is the principal site of estrogen formation in postmenopausal women. Aromatase activity as well as P450arom transcripts primarily reside in the stromal cell component of the adipose tissue. Studies designed to investigate whether increased local aromatase activity in breast adipose tissue influences the growth of breast cancers have yielded discrepant results. In an attempt to clarify this controversy, adipose tissue was obtained from the four breast quadrants at the time of mastectomy (n = 13) performed for removal of a tumor. Breast fat P450arom messenger RNA levels were quantified and compared between the four quadrants within each specimen using competitive polymerase chain reaction after reverse transcription in which 10 micrograms human adipose total RNA together with 1 pg rat complementary RNA (internal standard) were reverse transcribed and coamplified. In 9 out of 13 patients (69%), highest P450arom transcript levels colocalized to the quadrants bearing tumors. This correlation was statistically significant (P < 0.001). The regional distribution of P450arom transcripts in breast adipose tissue of disease-free individuals, obtained during reduction mammoplasty (control group, n = 9), did not favor any particular region of the breast. We also quantified by morphometry the histological components of the adipose tissue samples from each quadrant in mastectomy specimens. The distribution of stromal cells significantly correlated with the distribution of P450arom transcript levels, in that quadrants containing highest proportions of stromal cells matched to highest transcript levels (P < 0.01). Although the quadrants bearing tumors contained the highest percentage of stromal cells, this correlation was not statistically significant. The adipose tissue surrounding a breast tumor displays increased estrogen biosynthesis, which may promote tumor growth. It is further suggested that the distribution of stromal cell components in breast adipose tissue gives rise to locally elevated P450arom expression, which in turn may favor neoplastic development and growth in these predisposed areas of the breast. The correlation between the presence of a tumor and elevated P450arom levels in the proximal adipose tissue is independent of tumor size, node involvement, histological type or grade, estrogen/progesterone receptor status, DNA index, or S-phase fraction.

Adipose Tissue↗

In vivo transcript profiling and phylogenetic analysis identifies suppressor of cytokine signaling 2 as a direct signal transducer and activator of transcription 5b target in liver.

The GH-activated signal transducer and activator of transcription 5b (STAT5b) is an essential regulator of somatic growth. The transcriptional response to STAT5b in liver is poorly understood. We have combined microarray-based expression profiling and phylogenetic analysis of gene regulatory regions to study the interplay between STAT5b and GH in the regulation of hepatic gene expression. The acute transcriptional response to GH in vivo after a single pulse of GH was studied in the liver of hypophysectomized rats in the presence of either constitutively active or a dominant-negative STAT5b delivered by adenoviral gene transfer. Genes showing differential expression in these two situations were analyzed for the presence of STAT5b binding sites in promoter and intronic regions that are phylogenetically conserved between rats and humans. Using this approach, we showed that most rapid transcriptional effects of GH in the liver are not results of direct actions of STAT5b. In addition, we identified novel STAT5b cis regulatory elements in genes such as Frizzled-4, epithelial membrane protein-1, and the suppressor of cytokine signaling 2 (SOCS2). Detailed analysis of SOCS2 promoter demonstrated its direct transcriptional regulation by STAT5b upon GH stimulation. A novel response element was identified within the first intron of the human SOCS2 gene composed of an E-box followed by tandem STAT5b binding sites, both of which are required for full GH responsiveness. In summary, we demonstrate the power of combining transcript profiling with phylogenetic sequence analysis to define novel regulatory paradigms.

Animals↗

Requirement for the Drosophila COE transcription factor Collier in formation of an embryonic muscle: transcriptional response to notch signalling.

During Drosophila embryogenesis, mesodermal cells are recruited to form a stereotyped pattern of about 30 different larval muscles per hemisegment. The formation of this pattern is initiated by the specification of a special class of myoblasts, called founder cells, that are uniquely able to fuse with neighbouring myoblasts. We report here the role of the COE transcription factor Collier in the formation of a single muscle, muscle DA3([A])(DA4([T])). Col expression is first observed in two promuscular clusters (in segments A1-A7), the two corresponding progenitors and their progeny founder cells, but its transcription is maintained in only one of these four founder cells, the founder of muscle DA3([A]). This lineage-specific restriction depends on the asymmetric segregation of Numb during the progenitor cell division and involves the repression of col transcription by Notch signalling. In col mutant embryos, the DA3([A]) founder cells form but do not maintain col transcription and are unable to fuse with neighbouring myoblasts, leading to a loss-of-muscle DA3([A]) phenotype. In wild-type embryos, each of the DA3([A])-recruited myoblasts turns on col transcription, indicating that the conversion, by the DA3([A]) founder cell, of 'naive' myoblasts to express its distinctive pattern of gene expression involves activation of col itself. We find that muscles DA3([A]) and DO5([A]) (DA4([T]) and DO5([T])) derive from a common progenitor cell. Ectopic expression of Col is not sufficient, however, to switch the DO5([A]) to a DA3([A]) fate. Together these results lead us to propose that specification of the DA3([A]) muscle lineage requires both Col and at least one other transcription factor, supporting the hypothesis of a combinatorial code of muscle-specific gene regulation controlling the formation and diversification of individual somatic muscles.

Animals↗

More than 40,000 transcripts, including novel and noncoding transcripts, in mouse embryonic stem cells.

To study the transcriptome of embryonic stem cells, we used a new gene expression profiling method that can measure the expression levels of unknown and rarely expressed transcripts precisely. We detected a total of 33,136 signal peaks representing transcripts in mouse embryonic stem cells, E14. Subsequent random cloning of the peaks suggests that mouse embryonic stem cells express at least 40,000 transcripts, of which about 2,000 are still unknown. In addition, we identified 1,022 noncoding transcripts, several of which change depending on differentiation in gene expression. Our database provides a high-resolution expression profile of E14 cells and is applicable to other mouse embryonic stem cell analyses. It includes most transcription regulation factor-encoding genes and a significant number of unknown and noncoding transcripts.

Animals↗

Regulation of fatty acid synthase at transcriptional and post-transcriptional levels in rat liver.

The regulation of fatty acid synthase in rat liver was investigated at transcriptional and post-transcriptional levels. When rats were fasted for 3 days and refed on a high-carbohydrate diet, the amounts of FAS in liver cytosol began to increase at 12 hours and further increased until 48 hours. The amount of mRNA for FAS began to increase at 6 hours and reached to a maximum level at 12 hours, indicating that the expression of mRNA for FAS precedes the increase of FAS protein pool. After 12 hours the amounts of mRNA gradually decreased and remained at a much lowered level between 24 and 48 hours. The elevated amount of FAS mRNA reflected on the amount of FAS protein in the first 24 hours, but these two parameters were not paralleled thereafter, probably due to the changes in the translational efficiencies. The run-on transcriptional activity of FAS gene began to increase at 4 hours after refeeding a high-carbohydrate diet and further increased to reach a maximum level 25 fold of the initial level at 12 hours, followed by a 16 fold level between 24 and 48 hours. The elevation of run-on transcriptional activity of FAS gene preceded the increase of FAS mRNA in the liver cytosol by 2 hours, and a similar increasing pattern was observed until 12 hours. However, FAS mRNA concentration decreased gradually after 12 hours, while the transcriptional activity remained at a high level until 48 hours. The changes in FAS mRNA content in the cytosol of rat liver were closely related to the transcriptional activity of FAS gene in the early phase of induction, but another regulatory mechanism seems to operate in the decrease of mRNA after 12 hours.

Animals↗

Intergenic transcription is not required in Th2 cells to maintain histone acetylation and transcriptional permissiveness at the Il4-Il13 locus.

Noncoding RNA transcripts mapping to intergenic regions of the Il4-Il13 locus have been detected in Th2 cells harboring transcriptionally permissive Il4 and Il13 genes but not in Th1 cells where these genes are repressed. This correlation has given rise to the idea that intergenic transcription may be involved in maintaining the "open" chromatin structure of the Il4-Il13 locus in Th2 cells. We present evidence from real-time RT-PCR, nuclear run on, chromatin immunoprecipitation and 5,6-dichlorobenzimidazole 1-beta-D-ribofuranoside-mediated transcriptional inhibition analyses that argue against this hypothesis. Instead, our results are consistent with an alternative role for intergenic transcription in the maintenance of transcriptional silence in Th1-primed cells.

Acetylation↗

[Post-transcriptional regulation by 3'-untranslated region of transcripts].

Two different mechanisms have been well known to regulate the amounts of various transcripts in response to internal and external environmental stimuli. First, by binding of activated transcription factors to DNA regulatory regions including the cAMP response element, hormone response element, and activator protein-1 region upstream in various genes, the rate of transcription from DNA to mRNA is regulated. Secondly, the degradation of some mRNAs related to immune responses has been reported to be regulated by binding of RNA-binding proteins to adenylate uridylate-rich elements (AU-rich elements, AREs) located in the 3'-untranslated region (3'-UTR). The original study identifying the existence of a common regulatory nucleotide sequence in the 3'-UTR of inflammatory mediator transcripts pointed out that the AREs are characteristic of immune-related functional proteins. The number of transcripts containing AREs in the 3'-UTR has increased and several neuronal proteins including beta 2-adrenergic receptor, nerve growth factor, tyrosine hydroxylase, and nitric-oxide synthase II, have been reported to have AREs. We here reviewed the recent advances in the neuropsychopharmacological understanding of post-transcriptional regulation by RNA-binding protein and also pointed out the importance of this regulation in future studies using various stress paradigms.

3' Untranslated Regions↗

Prolonged IL-2 receptor alpha/CD25 expression after T cell activation via the adhesion molecules CD2 and CD28. Demonstration of combined transcriptional and post-transcriptional regulation.

The T cell adhesion molecule CD28 provides a costimulatory signal in combination with CD2 and CD3 mAb. CD28 regulates the expression of cytokines by T cells, not only IL-2, but also IL-1 alpha and CSF-1, usually synthesized by accessory cells. We have investigated the mechanisms through which CD28 modulates the expression of the IL-2R alpha chain. Whereas activation through CD2 or CD28 alone induced no or only low IL-2R alpha chain expression, activation through CD2 plus CD28 led to both a high and prolonged (greater than 14 days) cell surface and mRNA expression. In contrast, immobilized CD3 mAb-dependent activation induced a transient expression of the IL-2R alpha chain, which was neither further increased nor prolonged by CD28 costimulation. Upon CD2 plus CD28 stimulation, the half-lives of the two IL-2R alpha transcripts increased progressively between days 1 and 4, in contrast to each pathway alone. Whereas each activation pathway alone induced either no (CD2) or low (CD28) levels of IL-2R alpha gene transcription, the CD2 plus CD28 stimulation was associated with its increased transcription, which persisted at similar rates between 5 and 96 h post stimulation. The in vitro costimulation via the CD2 and CD28 molecules thus regulates the expression of the IL-2R alpha gene both at the transcriptional and post transcriptional levels. Our results therefore demonstrate a new immunoregulatory function of the CD28 molecule on IL-2R alpha expression, which, through its increased transcription rate and stabilization, could, together with high levels of cytokines secretion, be responsible for the prolonged T cell proliferation.

Antigens, CD↗