A distinct population of nonphagocytic and low level CD4+ null lymphocytes produce IFN-alpha after stimulation by herpes simplex virus-infected cells.

Human PBMC were stimulated for 6 h in vitro by HSV or Sendai virus (SV) and analyzed by flow cytometry. IFN-alpha producing cells (IPC) were identified through their content of IFN-alpha mRNA by in situ hybridization using a 35S-labeled IFN-alpha 2 cRNA probe. The IPC induced by HSV-infected WISH cells lacked capacity to adhere to and phagocytose latex particles. The induction of IFN-alpha by free infectious SV occurring in monocytes was abolished by phagocytosis of latex particles present in the cultures during the induction period. Such latex particles actually enhanced the IFN-alpha response induced by glutaraldehyde-fixed HSV- or SV-infected WISH cells or by free intact HSV. The HSV-induced IPC did not express the CD14 Ag expressed on monocytes. Cell sorting was performed on HSV-induced PBMC labeled with phycoerythrin-conjugated anti-CD3 and FITC-conjugated anti-CD4 mAb. A small population consisting of 1.4% of all PBMC, which was CD3- but expressed low but significant levels of CD4, contained the majority of the IPC with a 50-fold increase of their frequency. This cell population had a forward- and right-angle light scatter different from typical monocytes/macrophages. The results therefore further delineate IPC among PBMC into monocytes, being stimulated by viruses such as SV. Another distinct population of infrequent but highly efficient IPC, tentatively designated natural IFN-alpha producing cells, is activated by stimuli such as HSV.     

Characterization of interleukin 2-dependent cytotoxic T-cell clones. IV. Production of alpha, beta and gamma interferons and interleukin 2 by Lyt-2+ T cells

The production of alpha, beta and gamma interferons (IFN) and interleukin 2 (IL-2) by Lyt-2+-dependent cytotoxic T-cell lines/clones was investigated. Cloned and uncloned T-cell lines specific for H-2Dd or the unique RL male 1 leukemia antigen were studied. After infection with Sendai virus (SV) or Newcastle disease virus (NDV) all cell lines produced IFN-alpha and -beta. Induction of IFN-gamma was attempted with the mitogens Con A, PHA, PWM, SEA, and SEB, with poly(I:C), with antibodies Lyt-1.2, -2.2, and Thy-1.2, or with the target cells Meth A (H-2Dd+) and RL male 1. All mitogens were effective inducers. However, the antibodies and poly(I:C) were not. One uncloned RL male 1-specific cell line CTLL-RP, produced IFN-gamma after induction with RL male 1. Production of IFN-alpha, beta depended on IL-2, whereas production of IFN-gamma did not, although addition of highly purified IL-2 increased IFN-gamma production even in the absence of other inducers. Crude IL-2 inhibited the production of IFN-gamma but not IFN-alpha, beta. In response to mitogens, some T-cell clones also produced IL-2. The results demonstrate that Lyt-2+ cells can produce a broad spectrum of lymphokine activities after appropriate stimulation. Their availability now affords us the opportunity to study the regulation of lymphokine production at the clonal level.     

Degradation of STAT1 and STAT2 by the V proteins of simian virus 5 and human parainfluenza virus type 2, respectively: consequences for virus replication in the presence of alpha/beta and gamma interferons

Human cell lines were isolated that express the V protein of either simian virus 5 (SV5) or human parainfluenza virus type 2 (hPIV2); the cell lines were termed 2f/SV5-V and 2f/PIV2-V, respectively. STAT1 was not detectable in 2f/SV5-V cells, and the cells failed to signal in response to either alpha/beta interferons (IFN-alpha and IFN-beta, or IFN-alpha/beta) or gamma interferon (IFN-gamma). In contrast, STAT2 was absent from 2f/PIV2-V cells, and IFN-alpha/beta but not IFN-gamma signaling was blocked in these cells. Treatment of both 2f/SV5-V and 2f/PIV2-V cells with a proteasome inhibitor allowed the respective STAT levels to accumulate at rates similar to those seen in 2fTGH cells, indicating that the V proteins target the STATs for proteasomal degradation. Infection with SV5 can lead to a complete loss of both phosphorylated and nonphosphorylated forms of STAT1 by 6 h postinfection. Since the turnover of STAT1 in uninfected cells is longer than 24 h, we conclude that degradation of STAT1 is the main mechanism by which SV5 blocks interferon (IFN) signaling. Pretreatment of 2fTGH cells with IFN-alpha severely inhibited both SV5 and hPIV2 protein synthesis. However, and in marked contrast, pretreatment of 2fTGH cells with IFN-gamma had little obvious effect on SV5 protein synthesis but did significantly reduce the replication of hPIV2. Pretreament with IFN-alpha or IFN-gamma did not induce an antiviral state in 2f/SV5-V cells, indicating either that the induction of an antiviral state is completely dependent on STAT signaling or that the V protein interferes with other, STAT-independent cell signaling pathways that may be induced by IFNs. Even though SV5 blocked IFN signaling, the addition of exogenous IFN-alpha to the culture medium of 2fTGH cells 12 h after a low-multiplicity infection with SV5 significantly reduced the subsequent cell-to-cell spread of virus. The significance of the results in terms of the strategy that these viruses have evolved to circumvent the IFN response is discussed.     

In vivo and in vitro induction of (2'-5') oligoadenylate synthetase by human interferons in leukocytes from healthy donors and patients with renal cancer and hairy cell leukemia

The effect of in vivo administered interferon-alpha (IFN-alpha) on 2-5-oligoadenylate (A) synthetase activity of peripheral blood mononuclear cells (PBMC) was compared in patients with hairy cell leukemia and renal cell cancer. Basic levels of this enzyme varied from donor to donor, but mean levels were not significantly different in patients with renal cell cancer or hairy cell leukemia compared to healthy donors. After a single injection of 3 x 10(6) IU IFN, these basic levels rose 2- to 8-fold within 12-24 h post-injection and reverted to pretreatment levels after 48 h. The extent of this in vivo stimulation by IFN-alpha was similar in patients with hairy cell leukemia and renal cell carcinoma, and was correlated with down-regulation of IFN-alpha receptors. The in vitro effects of IFN-alpha, -beta and -gamma were compared after 18 h treatment with 10, 10(2) and 10(3) IU/ml of each IFN. Unlike IFN-alpha and -beta, IFN-gamma did not induce 2-5 A synthetase activity in either normal PBMC or hairy cells; these results were related to the effects of the three IFN on proliferative response of normal PBMC to phytohemagglutinin. Our data support the idea that 2-5 A synthetase activity is a marker of biological response to interferon treatment in human cancers.     

Modulation by interferons of the expression of monocyte complement genes.

Interferons-alpha, -beta and -gamma (IFNs-alpha, -beta and -gamma) stimulated the synthesis of the second complement component (C2), Factor B (B) and C1 inhibitor (C1-inh) by human monocytes in vitro. The degree of increase of the secretion rates of C2, B and C1-inh was dose-dependent and proportional to increases in the abundances of their respective mRNAs. IFN-gamma was the most effective at stimulating monocyte C1-inh synthesis, whereas IFN-alpha and IFN-beta were marginally more effective at stimulating monocyte C2 and B synthesis. Kinetic studies showed that the effect of the IFNs was rapid, with maximum stimulation occurring within 1-2 h for all three proteins. After the removal of IFNs from cultures the C1-inh mRNA abundance remained elevated for over 24 h in IFN-gamma-treated monocytes but returned to control levels within 8 h in IFN-alpha-treated and IFN-beta-treated monocytes. The abundances of C2 mRNA and B mRNA also returned to basal values within 8 h after removal of any of the three cytokines from the cultures. Both IFN-alpha and IFN-beta acted synergistically with IFN-gamma to stimulate synthesis of C1-inh and B. This synergistic effect only occurred when the cytokines were present in the cultures simultaneously. The effects of IFN-gamma plus IFN-alpha or IFN-beta on C2 synthesis appeared to be additive rather than synergistic. IFN-gamma inhibited synthesis of C3 by monocytes, but IFN-alpha and IFN-beta had no effect on the synthesis of this protein. Furthermore, none of the three cytokines had any effect on the expression of actin mRNA in monocytes.     

Fc gamma RIIa is expressed on natural IFN-alpha-producing cells (plasmacytoid dendritic cells) and is required for the IFN-alpha production induced by apoptotic cells combined with lupus IgG

An ongoing production of IFN-alpha may be of etiopathogenic significance in systemic lupus erythematosus (SLE). It may be due to the natural IFN-producing cells (NIPC), also termed plasmacytoid dendritic cells (PDC), activated by immune complexes that contain nucleic acids derived from apoptotic cells. We here examined the role of FcgammaR in the IFN-alpha production in vitro by PBMC induced by the combination of apoptotic U937 cells and autoantibody-containing IgG from SLE patients (SLE-IgG). The Fc portion of the SLE-IgG was essential to induce IFN-alpha production, because Fab fragments or F(ab')(2) were ineffective. Normal, especially heat-aggregated, IgG inhibited the IFN-alpha production, suggesting a role for FcgammaR on PBMC. Using blocking anti-FcgammaR Abs, the FcgammaRIIa,c (CD32) but not FcgammaRI or FcgammaRIII were shown to be involved in the IFN-alpha induction by apoptotic cells combined with SLE-IgG, but not by HSV or CpG DNA. In contrast, the action of all of these inducers was inhibited by the anti-FcgammaRIIa,b,c mAb AT10 or heat-aggregated IgG. Flow cytometric analysis revealed that approximately 50% of the BDCA-2-positive PBMC, i.e., NIPC/PDC, expressed low but significant levels of FcgammaRII, as did most of the actual IFN-alpha producers activated by HSV. RT-PCR applied to NIPC/PDC purified by FACS demonstrated expression of FcgammaRIIa, but not of FcgammaRIIb or FcgammaRIIc. We conclude that FcgammaRIIa on NIPC/PDC is involved in the activation of IFN-alpha production by interferogenic immune complexes, but may also mediate inhibitory signals. The FcgammaRIIa could therefore have a key function in NIPC/PDC and be a potential therapeutic target in SLE.     

Promotion of alpha/beta interferon induction during in vivo viral infection through alpha/beta interferon receptor/STAT1 system-dependent and -independent pathways

Viruses and viral components can be potent inducers of alpha/beta interferons (IFN-alpha/beta). In culture, IFN-alpha/beta prime for their own expression, in response to viruses, through interferon regulatory factor 7 (IRF-7) induction. The studies presented here evaluated the requirements for functional IFN receptors and the IFN signaling molecule STAT1 in IFN-alpha/beta induction during infections of mice with lymphocytic choriomeningitis virus (LCMV). At 24 h after infection, levels of induced IFN-alpha/beta in serum were reduced 90 to 95% in IFN-alpha/beta receptor-deficient (IFN-alpha/betaR(-/-)) and STAT1(-/-) mice compared to those in wild-type mice. However, at 48 h, these mice showed elevated expression in the serum whereas IFN-alpha/beta levels were still reduced >75% in IFN-alpha/betagammaR(-/-) mice even though the viral burden was heavy. Levels of IFN-beta, IFN-alpha4, and non-IFN-alpha4 subtype mRNA expression correlated with IFN-alpha/beta bioactivity, and all IFN-alpha/beta subtypes were coincidentally detectable. IRF-7 mRNA was induced under conditions of IFN-alpha/beta production, including late production in IFN-alpha/betaR(-/-) mice. These data demonstrate that the presence of the virus alone is not sufficient to induce IFN-alpha/beta during LCMV infection in vivo. Instead, autocrine amplification through the IFN-alpha/betaR is necessary for optimal induction. In the absence of a functional IFN-alpha/betaR, however, alternative mechanisms, independent of STAT1 but requiring a functional IFN-gammaR, take over.     

Effect of interferon on replication of herpes simplex virus types 1 and 2 in human macrophages.

Macrophages derived from human peripheral blood and cultured for 1 week were permissive for the replication of herpes simplex virus (HSV) types 1 and 2. Low titers of interferon (IFN) were produced after virus infection. The yield of infectious virions was reduced by pretreatment of cells with natural and recombinant IFN-alpha and natural IFN-beta. Recombinant and natural IFN-gamma exhibited very low antiviral activity. Treatment of cells with IFN-gamma mixed with IFN-alpha or with IFN-beta did not result in a synergistic inhibition of virus yield. We studied the synthesis of HSV type 1- and HSV type 2-coded proteins in macrophages treated with IFN-beta. Induction of the HSV beta-protein DNA polymerase was strongly inhibited in IFN-treated cells in a dose-dependent manner. As shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, other beta- and gamma-proteins of HSV were inhibited as well. Immunofluorescence studies revealed a strong inhibition of the expression of immediate early alpha-protein ICP4. The results indicate that IFN acts early during the viral replication cycle to inhibit the synthesis of HSV alpha- and beta-proteins.     

Mutational analysis of regulation of MHC and anti-viral genes.

CTL-mediated selection for loss of expression of Mta by H-2-heterozygous SV40-transformed mouse fibroblasts (line 24SV) produced an unusual phenotypic class of maternally transmitted Ag negative mutants defective in both MHC expression and in anti-viral activity. Severely reduced surface expression of class I MHC Ag from multiple loci of both haplotypes correlated with low levels of MHC H chain and beta 2-microglobulin mRNA. Inasmuch as IFN can up-regulate class I expression and some fibroblasts elaborate autocrine IFN-beta, we examined whether IFN could restore wild-type expression of class I MHC Ag. However, IFN could not restore wild-type expression. Moreover, the fold-increases in class I Ag and mRNA expression were significantly reduced in mutant cells compared to wild-type cells. These results suggested that the mutants might have generalized defects in IFN response. Inasmuch as the induction of an anti-viral state is a hallmark of IFN responses, we exposed cells to IFN-alpha, -beta, or -gamma and challenged with virus. 24SV cells, exposed to any of the three IFNs, were completely protected from destruction by vesicular stomatitis, mengovirus or respiratory syncytial viruses. In contrast, MHC and anti-viral defective mutants could not be protected from virus-induced lysis by any IFN. Somatic cell hybridization analyses indicated that both basal MHC and IFN-inducible phenotypes were recessive to wild-type, and that a trans-acting regulatory factor required for basal MHC expression is defectively expressed in the mutants. Such a factor may integrate the organismal response to virus infection, encompassing both immune and nonimmune anti-viral responses.     

Activity of the simian virus 40 early promoter-enhancer in herpes simplex virus type 1 vectors is dependent on its position, the infected cell type, and the presence of Vmw175.

We have studied some of the parameters governing the expression of a foreign promoter-reporter gene construct incorporated into herpes simplex virus (HSV) type 1. These include the genetic background of the parental virus, the site of transgene insertion within the HSV genome, and the infected cell type. The genetic background of the vector constructs denoted delta 3 was an HSV type 1 mutant deleted for nearly the entire coding portion of Vmw175 (ICP4), the product of the essential immediate-early gene IE3. For vectors denoted +, the IE3 deletion had been repaired by marker rescue. We used as a reporter gene the bacterial chloramphenicol acetyltransferase (CAT) gene, driven by the simian virus 40 (SV40) early promoter and enhancer region. The SV40-cat hybrid gene was inserted either into the HSV thymidine kinase (TK) locus to create the vectors TKScat delta 3 and TKScat+ or into an intergenic site within the BamHI z fragment of the short unique portion of the viral genome to create the vectors GScat delta 3 and GScat+. In Vero and BHK cells infected with TKScat delta 3, CAT activity was first detected at 10 h postinfection and continued to accumulate until 36 h postinfection. In cells of primate origin infected with the replication-competent vector TKScat+, or in primate cells which complement the IE3 deficiency and which were infected with TKScat delta 3, CAT activity was significantly lower than in cells of rodent origin. However, levels of CAT were increased in the presence of cycloheximide, suggesting that the low production of CAT in primate cells was due to repression of SV40-cat hybrid gene expression. In contrast with results with TKScat delta 3 and TKScat+, CAT activity was not detectable in any of the tested cell types infected with GScat delta 3 or GScat+ except under conditions of cycloheximide reversal. These results show that while HSV gene products expressed in the presence of Vmw175 inhibited SV40-cat expression in the tk locus in a cell-type-specific manner, HSV gene products expressed in the presence or absence of Vmw175 inhibited SV40-cat expression in the BamHI z locus independently of cell type.     

Different binding of human interferon alpha 1 and alpha 2 to common receptors on human and bovine cells. Studies with recombination interferons produced in Escherichia coli

Minicells from Escherichia coli DS410 harboring cDNA for human interferon (IFN) alpha 1 or alpha 2 were metabolically labeled with [3H]leucine and the radioactive IFN was purified to homogeneity by immune precipitation with anti-IFN-alpha serum. These preparations of radioactive IFN-alpha 1 and -alpha 2 were used to study the binding on two human (FL and Daudi) and one bovine (MDBK) cell lines. IFN-alpha 2 specifically bound well to both human and bovine cells, while IFN-alpha 1 bound very poorly to human cells but well to bovine cells. Specific binding of radioactive IFN-alpha 2 to these cell lines was completely inhibited by not only nonradioactive IFN-alpha 2 but also IFN-alpha 1, and binding of IFN-alpha 1 to bovine cell was also competed by IFN-alpha 2 as well as IFN-alpha 1, indicating that the receptors for both IFNs are identical. However, 50-100-fold (on human cells) or 4-fold (on bovine cell) more nonradioactive IFN-alpha 1 than -alpha 2 was required to inhibit the binding of radioactive IFN-alpha 2 to the receptors. Scatchard analysis showed that IFN-alpha 1 and -alpha 2 bind to the receptors on human cells with an apparent Kd of greater than 6 X 10(-10) and 3 X 10(-11) M, respectively, while on bovine cells with a Kd of 4.2 X 10(-11) and 1.6 X 10(-11) M, respectively. These results show that the different target cell specificity of IFN-alpha 1 and -alpha 2 in regard to antiviral activity (Streuli, M., Hall, A., Boll, W., Stewart, W. E., II, Nagata, S., and Weissmann, C. (1981) Proc. Natl. Acad. Sci. U. S. A. 78, 2848-2852) is due to the different binding activity of IFN-alpha molecules to their common receptors.     

Activation of caspase-3 by interferon alpha causes interleukin-16 secretion but fails to modulate activation induced cell death

Interferon alpha (IFN-alpha) is the mainstay in the treatment of chronic hepatitis C virus (HCV) infection. Interleukin-16 (IL-16) attracts CD4+ cells to sites of inflammation and plays a role in the interaction of dendritic cells, T cells and B cells. In this study, we show that IFN-alpha itself induces IL-16 secretion by peripheral blood lymphocytes (PBL) and enhances IL-16 secretion by anti-CD3 stimulated PBL. Pro-IL-16 is cleaved into its active form by caspase-3. IFN-alpha increases caspase-3 mRNA levels in activated T cells (ATC), as shown by Northern blot analysis, whereas IL-16 mRNA levels are not affected by IFN-alpha. IL-16 secretion into culture supernatants correlates tightly with intracellular caspase-3 activity in ATC. In our experiments addition of specific caspase inhibitors did not reduce the proportion of ATC undergoing Fas-mediated cell death, but completely blocked IFN-alpha-induced IL-16 secretion into culture supernatants. In conclusion, our results suggest that IFN-alpha activates caspase-3, thereby increasing secretion of IL-16, whereas IFN-alpha-enhanced Fas-mediated cell death in ATC is not caspase-dependent.     

Mouse genotype affects inducible expression of cytokine genes.

The levels of circulating IFN in mice infected with Newcastle disease virus (NDV) are regulated by the If-1 locus. In this study we show that in NDV-infected C57BL/6 mice, which carry the If-1h allele and produce high levels of IFN, high levels of both IFN-alpha and -beta mRNA can be detected in the spleen. In contrast, only very low levels of IFN mRNA could be detected in spleens of infected BALB/c mice containing the If-1l allele and producing low levels of IFN or in B6.C-H28c mice that are congenic for the If-1l allele. The relative levels of all individual IFN-alpha 1, alpha 4, and alpha 6 mRNA in spleens of infected BALB/c were lower than in spleens of infected C57BL/6 mice, indicating that the If-1 locus affects the expression of all IFN-alpha subtypes and is not associated with the deletion or inactivation of a specific IFN gene. The relative levels of IFN regulatory factor-1 mRNA in infected mice carrying the If-1l and If-1h loci were comparable, suggesting that the If-1 regulation is not associated with the altered expression of the IFN regulatory factor-1 gene. Quantitative difference in the expression of IFN-alpha and -beta genes was also observed in in vitro-infected peritoneal macrophages isolated from either C57BL/6 or BALB/c mice. A surprise finding was that the If-1 locus also affected the NDV-induced expression of two other cytokine genes, TNF-alpha and IL-6. Priming of the macrophage cultures with murine IFN enhanced the expression of all cytokine genes, and the relative levels of IFN, TNF-alpha, and IL-6 mRNA induced by NDV in macrophages derived from C57BL/6 and BALB/c mice were comparable. We propose that the If-1 locus affects the early stages of a signal transduction pathway which are common to the virus-mediated induction of IFN, TNF-alpha, and IL-6 genes.     

Human cytokines, tumor necrosis factor, and interferons: gene cloning, animal studies, and clinical trials

Presented is a comprehensive program designed to isolate human cytokine genes and investigate their relative induction, and to analyze cytokine activities in cell culture, animal tumor models, and human clinical trials. Human cytokine cDNAs have been isolated from a cDNA library made from normal human peripheral blood leukocytes (PBLs) treated with Sendai virus and the relative induction of tumor necrosis factor (TNF), alpha and gamma interferons (IFN-alpha, IFN-gamma), and interleukin-1 beta IL-1 beta) genes has been analyzed. In the Sendai virus-induced PBL system, IL-1 beta mRNA was shown to be approximately twofold higher than TNF or IFN-alpha mRNA whereas IFN-gamma mRNA was 50-100-fold lower than TNF or IFN-alpha mRNA. The cytotoxic activity of TNF was analyzed on several cell lines and IFN-alpha and IFN-gamma were shown to potentiate TNF cytotoxicity about 2-200-fold depending on cell lines. The LD50 for recombinant TNF in BALB/c mice was determined to be 6 X 10(7) U/kg and the therapeutic dose of recombinant TNF in sarcoma 180 bearing BALB/c mice was 3 X 10(5) U/kg, indicating a wide therapetic index. Phase I clinical trials of recombinant TNF given I.V. indicated a tolerated dose of 150,000 U/kg with biphasic half-life (T-1/2) of 2 and 31 min following TNF injection. Phase II trials of TNF and trials of TNF combined with IFN-alpha are in progress. These studies indicate that cytokines such as TNF and IFN-alpha are subject to similar induction systems, potentiate each other's activities, and can be tolerated at specific doses for potential therapeutic use.     

A human cytomegalovirus function inhibits replication of herpes simplex virus.

Human embryonic lung (HEL) cells infected with human cytomegalovirus (HCMV) restricted the replication of herpes simplex virus type 1 (HSV-1). A delay in HSV replication of 15 h as well as a consistent, almost 3 log inhibition of HSV replication in HCMV-infected cell cultures harvested 24 to 72 h after superinfection were observed compared with controls infected with HSV alone. Treatment of HCMV-infected HEL cells with cycloheximide (100 micrograms/ml) for 3 or 24 h, conditions known to result in accumulation of HCMV immediate-early and early mRNA, was demonstrated effective in blocking HCMV protein synthesis, as shown by immunoprecipitation with HCMV antibody-positive polyvalent serum. Cycloheximide treatment of HCMV-infected HEL cells and removal of the cycloheximide block before superinfection inhibited HSV-1 replication more efficiently than non-drug-treated superinfected controls. HCMV DNA-negative temperature-sensitive mutants restricted HSV as efficiently as wild-type HCMV suggesting that immediate-early and/or early events which occur before viral DNA synthesis are sufficient for inhibition of HSV. Inhibition of HSV-1 in HCMV-infected HEL cells was unaffected by elevated temperature (40.5 degrees C). However, prior UV irradiation of HCMV removed the block to HSV replication, demonstrating the requirement for an active HCMV genome. HSV-2 replication was similarly inhibited in HCMV-infected HEL cells. However, replication of adenovirus, another DNA virus, was not restricted in these cells under the same conditions. Superinfection of HCMV-infected HEL cells with HSV-1 labeled with [3H]thymidine provided evidence that the labeled virus could penetrate to the nucleus of cells after superinfection. Evidence for penetration of superinfecting HSV into HCMV-infected cells was also provided by blot hybridization of HSV DNA synthesized in cells infected with HSV alone versus superinfected cell cultures at 0 and 48 h after superinfection. In addition, superinfection with vesicular stomatitis virus ruled out a role for interferon in restriction of HSV replication in this system.     

Modulation of tubulin mRNA levels by interferon in human lymphoblastoid cells.

Blot hybridization with labeled tubulin cDNA showed that treatment of Ramos cells, a human cell line of lymphoblastoid origin, with either alpha or beta interferon (IFN) induced a marked increase in the amount of tubulin mRNA sequences. The level of tubulin mRNA sequences increased rapidly after exposure of cells to IFN-alpha and reached a maximum after 1 h of treatment, which was four times the control level. Treatment with IFN-beta induced a maximal increase after 4 h; the amount of tubulin mRNA sequences was seven times higher than the control level. The mRNA extracted from IFN-treated and nontreated cells was translated in vitro in a reticulocyte lysate cell-free system containing [35S]methionine. Electrophoretic analysis of the labeled cell-free products showed an increase in the amount of translatable tubulin mRNA that parallels the time course of induction of tubulin mRNA sequences. Two-dimensional gel electrophoresis of the labeled protein products directed by mRNA indicates that IFN caused a more pronounced increase in the level of alpha-tubulin than beta-tubulin mRNA. Treatment with colchicine, which disrupts the cell microtubules, caused a marked decrease in the tubulin mRNA content. Concomitant treatment of the cells with colchicine and IFN abolished the interferon-dependent induction of tubulin mRNA.