Biomedical subjects

T Inukai

Publications and source records attributed to T Inukai.

At least 55 records · Page 3Linked to original sources

PubMed2000

Estrogen markedly increases LDL-receptor activity in hypercholesterolemic patients.

We investigated the change in low-density lipoprotein receptor activity following the administration of estrogen to patients with hypercholesterolemia (HC). Studies were conducted in 16 patients with HC (total cholesterol (TC)>220 mg/dL) and 133 healthy control subjects. LDL-R activity was measured by fluorocytometry. Three of sixteen patients with HC showed low LDL-R activity below 80% together with extremely high serum levels of TC and LDL cholesterol (LDL-C). LDL-R activity showed an inverse correlation with serum levels of TC and LDL-C (p<0.05, respectively). Two women with initially low LDL-R activity who showed a marked increase in LDL-R activity exhibited a normalized activity at four and eight weeks after estriol administration, proportional to the reduction in serum levels of TC and LDL-C. One man with an initially extremely low LDL-R activity showed no abnormality at the sites of exons 7, 14, 17 and intron 12 by the PCR-DGGE method, which are common sites for point-mutations of LDL-R among Japanese patients with HC. Estrogen reduced serum levels of TC and LDL-C in patients with HC, at least in part, via an increase in the LDL-R activity of patients with HC and an initially low LDL-R activity.

Adult↗

PubMed1999

Two candidate downstream target genes for E2A-HLF.

The E2A-HLF fusion gene, formed by the t(17;19)(q22;p13) chromosomal translocation, is thought to drive the leukemic transformation of early B-cell precursors by repressing an evolutionarily conserved apoptotic pathway. To test this hypothesis, we sought to identify downstream targets of E2A-HLF in t(17;19)+ pro-B leukemia cells (UOC-B1) that had been transfected with a zinc-inducible vector encoding a dominant-negative suppressor (E2A-HLF[dn]) of the oncoprotein. Representational difference analysis of mRNAs from E2A-HLF(dn)+ UOC-B1 cells grown with (E2A-HLF inactive) or without (E2A-HLF active) the addition of zinc yielded several differentially expressed cDNA fragments that were individually subcloned. Two of the clones, designated F-5 and G-4, hybridized with mRNAs that were upregulated by E2A-HLF. Levels of both transcripts declined sharply within 8 to 12 hours after suppression of E2A-HLF DNA-binding activity, becoming undetectable after 96 hours. The F-5 cDNA was identified as a portion of ANNEXIN VIII, whose product was expressed in promyelocytic leukemia cells and UOC-B1 cells, but not in other leukemic cell lines. A novel full-length cDNA cloned with the G-4 fragment encoded a protein that we have named SRPUL (sushi-repeat protein upregulated in leukemia). It is normally expressed in heart, ovary, and placenta, but could not be detected in leukemic cell lines other than UOC-B1. Neither protein prevented apoptosis in interleukin-3-dependent murine pro-B cells, suggesting that they have paraneoplastic roles in leukemias that express E2A-HLF, perhaps in the disseminated intravascular coagulopathy and hypercalcemia that characterize these cases.

Amino Acid Sequence↗

PubMed1999

Running exercise increases tumor necrosis factor-alpha secreting from mesenteric fat in insulin-resistant rats.

Tumor necrosis factor-alpha (TNF-alpha) is an important mediator of insulin resistance in obese subjects, through its overexpression in fat tissue. However, how exercise can modify the expression of TNF-alpha is controversial. We examined TNF-alpha in adipose tissue using an animal model of insulin resistance that was produced by feeding rats a diet high in sucrose. The rats were allocated to one of three groups: those receiving a starch-based diet (control group): those fed a high-sucrose diet (sucrose-fed group): and those fed a high-sucrose diet and given wheel exercise (exercised group). The animals were allowed to eat and drink ad lib for 4 or 12 weeks (4 wk: control n=7, sucrose-fed n=7, exercised n=10; 12 wk: control n=5, sucrose-fed n=5, exercised n=9). The voluntary wheel exercise was initiated with the feeding of the high-sucrose diet. The rats in the exercise groups ran 15 +/- 3 km/week. We showed that 12-week voluntary running exercise significantly (P<0.05) increased both TNF-alpha protein (5-fold) and mRNA (1.4 fold) in the mesenteric fat of insulin-resistant rats compared to non-exercised sucrose-fed mice. Accordingly, in exercised group, plasma glucose (124 +/- 9 mEq/L vs 141 +/- 11 mEq/L). and free fatty acid (0.98 +/- 0.07 mEq/L vs 1.4 +/- 0.05 mEq/L) concentrating in portal vein blood were reduced compared to sucrose-fed group. The amounts of fatty tissue both in mesenteric and subcutaneous tissues were significantly (P<0.05) decreased through running exercise. We consider that up-regulation of TNF-alpha in mesenteric fat may be a compensatory mechanism for the reduction of fatty acid in adipose tissues and this change could control metabolic homeostasis during exercise to modulate a hyperinsulinemic state.

Adipose Tissue↗

PubMed1999

Preperitoneal fat deposition estimated by ultrasonography in patients with non-insulin-dependent diabetes mellitus.

UNLABELLED: Preperitoneal fat is an indicator of visceral fat deposition, which is closely related to atherosclerosis and coronary heart disease in obese patients. We assessed the relationship of preperitoneal fat deposition and various clinical characteristics in 90 patients with non-insulin-dependent diabetes mellitus (NIDDM). Preperitoneal and subcutaneous fat deposition were measured by ultrasonography. In both the male and female diabetics, preperitoneal fat levels were significantly higher than in age-matched healthy subjects. We also determined blood pressures, fasting plasma glucose, glycosylated hemoglobin A1c, serum lipids, fasting immunoreactive insulin (FIRI), daily urinary C-peptide (CPR), serum leptin, urinary albumin excretion and body mass index (BMI). Of these parameters, BMI, FIRI, leptin and daily urinary CPR were positively correlated with preperitoneal fat deposition. Patients with diet therapy alone showed significantly higher preperitoneal fat levels than those receiving insulin therapy. In female, patients with increased preperitoneal fat showed higher prevalence of hypertension than those with decreased fat. Macroalbuminuric patients had a lower preperitoneal fat than microalbuminuric and normoalbuminuric patients. Patients with proliferative retinopathy exhibited lower preperitoneal fat than did those without retinopathy. Preperitoneal fat levels were positively correlated with motor or sensory nerve conduction velocity. CONCLUSION: The present findings suggest that in NIDDM patients, increased preperitoneal fat deposition is closely associated with obesity, hypertension and hyperinsulinemia, and negatively modulates diabetic microangiopathy including nephropathy, retinopathy and neuropathy.

Adipose Tissue↗

PubMed1999

SLUG, a ces-1-related zinc finger transcription factor gene with antiapoptotic activity, is a downstream target of the E2A-HLF oncoprotein.

The E2A-HLF fusion gene transforms human pro-B lymphocytes by interfering with an early step in apoptotic signaling. In a search for E2A-HLF-responsive genes, we identified a zinc finger transcription factor, SLUG, whose product belongs to the Snail family of developmental regulatory proteins. Importantly, SLUG bears close homology to the CES-1 protein of C. elegans, which acts downstream of CES-2 in a neuron-specific cell death pathway. Consistent with the postulated role of CES-1 as an antiapoptotic transcription factor, SLUG was nearly as active as Bcl-2 or Bcl-xL in promoting the survival of IL-3-dependent murine pro-B cells deprived of the cytokine. We conclude that SLUG is an evolutionarily conserved transcriptional repressor whose activation by E2A-HLF promotes the aberrant survival and eventual malignant transformation of mammalian pro-B cells otherwise slated for apoptotic death.

Animals↗

PubMed1999

Facile enzymatic conversion of lactose into lacto-N-tetraose and lacto-N-neotetraose.

Lacto-N-tetraose (Galbeta1 -3GlcNAcbeta1-3Galbeta1-4Glc, LNT) and lacto-N-neotetraose (Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc, LNnT) were enzymatically synthesized by consecutive additions of GlcNAc and Gal residues to lactose. Lacto-N-triose II (GlcNAcbeta1-3Galbeta1-4Glc) was prepared first by the transfer of GlcNAc from UDP-GlcNAc to lactose by beta-1,3-N-acetylglucosaminyltransferase from bovine serum. The resulting lacto-N-triose II was converted into LNT and LNnT utilizing two kinds of beta-D-galactosidase-mediated transglycosylations. Thus, beta-D-galactosidase from Bacillus circulans ATCC31382 induced regioselective galactosyl transfer from o-nitrophenyl beta-D-galactoside to the OH-3" position of lacto-N-triose II, and commercially available beta-D-galactosidase from B. circulans to the OH-4" position of lacto-N-triose II. These convenient processes are suitable for large-scale preparations of LNT and LNnT. As another method, LNT was directly synthesized from lactose as an initial substance, utilizing lacto-N-biosidase (Aureobacterium sp. L-101)-mediated transglycosylation with Galbeta1-3GlcNAcbeta-pNP donor.

Animals↗

PubMed1999

The KOR-SA3544 antigen predominantly expressed on the surface of Philadelphia chromosome-positive acute lymphoblastic leukemia cells is nonspecific cross-reacting antigen-50/90 (CD66c) and invariably expressed in cytoplasm of human leukemia cells.

We previously reported a novel monoclonal antibody KOR-SA3544 which predominantly reacted with a surface antigen (sSA3544) expressed on Philadelphia chromosome (Ph1)-positive acute lymphoblastic leukemia (ALL). In the present study, we demonstrate that the antibody specifically recognized nonspecific cross-reacting antigen (NCA)-50/90 (CD66c), one of the carcinoembryonic antigen (CEA)-related glycoproteins encoded by a member of the CEA gene family. In addition, we show that the SA3544 antigen (NCA-50/90) was invariably expressed in cytoplasm of all of the human leukemic cell lines examined (sSA3544-positive B-lymphoid two, sSA3544-negative T or B-lymphoid and non-lymphoid 24) regardless of the presence or absence of surface expression of this antigen. Immunoelectromicroscopic examination revealed that the cytoplasmic antigen was mainly present in granules in sSA3544-positive leukemia cells, whereas it was diffusely present in cytosol in sSA3544-negative leukemia cells. Thus, among members of the CEA family, NCA-50/90 was first demonstrated to be expressed not only on the surface of some leukemia cells, but also in cytoplasm of various types of leukemia cells.

Antibodies, Monoclonal↗

PubMed1999

p16/MTS1/INK4A gene is frequently inactivated by hypermethylation in childhood acute lymphoblastic leukemia with 11q23 translocation.

The p16 gene encoding a specific inhibitor of cyclin-dependent kinases 4 and 6 has been reported to be inactivated at a variety of rates in malignant tumors. We studied frequency and mechanism of inactivation of the p16 gene in various types of childhood acute lymphoblastic leukemia (ALL) using 36 leukemic cell lines established from children (B precursor-ALL, 28; B-ALL/Burkitt's lymphoma, 3; and T-ALL, 5). On Southern blot, homozygous deletions or hemizygous deletions with rearrangement were detected in 14 cell lines. The expression of p16 protein was not observed on Western blot in 18 of 22 cell lines with intact p16 gene, but induced in 16 cell lines after treatment with the demethylating agent, indicating the silencing of the p16 gene by hypermethylation. Of note, the p16 gene was inactivated by hypermethylation of the 5' CpG island in nine of nine cell lines with 11q23 translocation, but was restored with the treatment of the demethylating agent. Partial methylation of the p16 gene was also demonstrated in three of eight primary leukemia samples with this translocation, suggesting that the p16 gene inactivation by hypermethylation might play a role in the leukemogenesis and disease progression of ALL with 11q23 translocation.

Blotting, Southern↗

PubMed1999

Relation between the serum level of C-peptide and risk factors for coronary heart disease and diabetic microangiopathy in patients with type-2 diabetes mellitus.

Syndrome X is used to describe a constellation of factors that lead to coronary heart disease (CHD): hypertension, hyperinsulinemia, impaired glucose tolerance, and an abnormality in lipid metabolism. We investigated the relationship between serum levels of C-peptide immunoreactivity (CPR) and diabetic complications in 256 patients with type-2 diabetes mellitus. The serum level of CPR was measured by radioimmunoassay (RIA). Diabetic patients were divided into 3 groups according to the serum level of CPR as follows: low CPR (n = 19, <0.7 ng/ml), normal CPR (n = 174, 0.7 to 2.2 ng/ml) and high CPR (n = 63, >2.2 ng/ml). The body mass index (BMI) and the serum level of triglycerides were significantly higher in the high CPR group (P < 0.05, respectively) compared with normal CPR group. The prevalence of hypertension was significantly higher in the high CPR group than in the other 2 groups (low CPR: 16%, normal CPR: 28%, high CPR: 38%). The frequency of the number of patients receiving insulin therapy was greater in the low CPR group than in the other 2 groups, (low CPR: 58%, normal CPR: 15%, high CPR: 11%). The serum CPR level was significantly lower in patients with than without proliferative retinopathy or macroalbuminuria. Our conclusion is that the present data suggest that an increased serum level of CPR is associated with obesity, elevated serum triglycerides, and hypertension in patients with type-2 diabetes mellitus. A low CPR level leading to hyperglycemia is associated with the progression of diabetic microangiopathies, such as retinopathy and nephropathy.

Albuminuria↗

PubMed1999

Reduction of leptin precedes fat loss from running exercise in insulin-resistant rats.

Serum concentrations of leptin, a hormone secreted into the circulation by adipocytes, correlate with body mass index. Circulating of leptin is thought to signal the brain in patients with hyperinsulinemia, a condition reported to be preventable and testable by exercise training. In the present experiments, sucrose-fed rats had reduced concentrations of leptin in portal venous blood after 4 weeks of nonforced wheel-running exercise (1.1 +/- 0.1 vs. 6.2 +/- 1.8 ng/mL, in nonexercised sucrose-fed rats, P < 0.05). Mesenteric and subcutaneous fat stores were similar between groups. After 12 weeks of exercise, portal vein levels of leptin concentrations (5.2 +/- 2.1 vs. 9.9 +/- 0.8 ng/mL, P < 0.05) and mesenteric and subcutaneous fat all were reduced in the exercise group. These results suggest that short-term running exercise reduces circulating leptin before any reduction of adipose mass, and this reduction in the concentration of leptin available to its receptors has beneficial effects on the metabolism of fat and carbohydrates.

Adipose Tissue↗

PubMed1999

Thyroid hormone modulates insulin-like growth factor-I(IGF-I) and IGF-binding protein-3, without mediation by growth hormone, in patients with autoimmune thyroid diseases.

The expression and synthesis of insulin-like growth factor-1 (IGF-I) and IGF-binding protein-3 (IGFBP-3) are regulated by various hormones and nutritional conditions. We evaluated the effects of thyroid hormones on serum levels of IGF-I and IGFBP-3 levels in patients with autoimmune thyroid diseases including 54 patients with Graves' disease and 17 patients with Hashimoto's thyroiditis, and in 32 healthy age-matched control subjects. Patients were subdivided into hyperthyroid, euthyroid and hypothyroid groups that were untreated, or were treated with methylmercaptoimidazole (MMI) or L-thyroxine (L-T4). Serum levels of growth hormone (GH), IGF-I and IGFBP-3 were determined by radioimmunoassay. Serum GH levels did not differ significantly between the hyperthyroid and the age-matched euthyroid patients with Graves' disease. The serum levels of IGF-I and IGFBP-3 showed a significant positive correlation in the patients (R=0.616, P<0.001). The levels of both IGF-I and IFGBP-3 were significantly higher in the hyperthyroid patients with Graves' disease or in those with Hashimoto's thyroiditis induced by excess L-T4 administration than in control subjects. Patients with hypothyroid Graves' disease induced by the excess administration of MMI showed significantly lower IGFBP-3 levels as compared to those in healthy controls (P<0.05). Levels of IGFBP-3, but not IGF-I levels, showed a significant positive correlation with the levels of free T4 and free T3. In Graves' disease, levels of TPOAb, but not of TRAb, showed a significant positive correlation with IGFBP-3. We conclude that in patients with autoimmune thyroid diseases, thyroid hormone modulates the synthesis and/or the secretion of IGF-I and IGFBP-3, and this function is not mediated by GH.

Adult↗

PubMed1999

Two distinct interleukin-3-mediated signal pathways, Ras-NFIL3 (E4BP4) and Bcl-xL, regulate the survival of murine pro-B lymphocytes.

Hematopoietic cells require cytokine-initiated signals for survival as well as proliferation. The pathways that transduce these signals, ensuring timely regulation of cell fate genes, remain largely undefined. The NFIL3 (E4BP4) transcription factor, Bcl-xL, and constitutively active mutants of components in Ras signal transduction pathways have been identified as key regulation proteins affecting murine interleukin-3 (IL-3)-dependent cell survival. Here we show that expression of NFIL3 is regulated by oncogenic Ras mutants through both the Raf-mitogen-activated protein kinase and phosphatidylinositol 3-kinase pathways. NFIL3 inhibits apoptosis without affecting Bcl-xL expression. By contrast, Bcl-xL levels are regulated through the membrane proximal portion in the cytoplasmic domain of the receptor (betac chain), which is shared by IL-3 and granulocyte-macrophage colony-stimulating factor. Activation of either pathway alone is insufficient to ensure cell survival, indicating that multiple independent signal transduction pathways mediate the survival of developing B-lymphoid cells.

Animals↗

PubMed1999

[Anti-thyroid peroxidase antibody].

Anti-thyroid peroxidase antibody(TPOAb) in patients with autoimmune thyroid diseases was identified with anti-microsomal antibody. Highly sensitive assays of TPOAb using radioimmunoassays and enzyme immunoassays have been developed following a hemagglutination assay technique. TPOAb is present in the serum of almost all patients with Hashimoto's thyroiditis, in more than 70% of those with Graves' disease, and, to a variable degree, in patients with nonthyroid autoimmune diseases and some normal subjects. There is a good correlation between the degree of lymphocytic infiltration of the thyroid gland and the titer of TPOAb. Postpartum thyroiditis, usually a transient disorder, occurs with high frequency in women who have TPOAb. TPOAb, in addition to anti-thyroglobulin antibody, was a clinically useful marker for defining autoimmune thyroid diseases and for detecting an underlying autoimmune process in the thyroid, however, its biological importance remains to be clarified at the present time.

Autoantibodies↗

PubMed1998

The chimeric E2A-HLF transcription factor abrogates p53-induced apoptosis in myeloid leukemia cells.

Leukemic lymphoblasts expressing the E2A-HLF oncoprotein possess wild-type p53 genes, but do not undergo apoptosis in response to DNA damage. Experimentally, E2A-HLF prevents apoptosis due to growth factor deprivation or gamma-irradiation in interleukin-3 (IL-3)-dependent murine pro-B cells. To directly test the chimeric protein's ability to abrogate p53-mediated cell death, we used mouse myeloid leukemia cells (M1p53tsval) that constitutively express a temperature-sensitive (ts) mutant p53 gene and undergo apoptosis when p53 assumes an active wild-type configuration. This effect is blocked by treatment with IL-6, which allows the cells to survive in culture despite wild-type p53 activation. We introduced E2A-HLF into M1p53tsval cells and found that they were resistant to p53-mediated apoptosis and that E2A-HLF effectively substituted for the survival functions of IL-6. The expression of p53-responsive genes such as p21 and Bax was upregulated normally, suggesting that E2A-HLF acts downstream of p53 to block execution of the p53-induced apoptotic program. NFIL3, a growth factor-regulated bZIP protein that binds to the same DNA-consensus site as E2A-HLF, delays apoptosis in IL-3-dependent pro-B cells deprived of growth factor. By contrast, in the present study, enforced expression of NFIL3 failed to protect M1p53tsval cells from p53-dependent apoptosis and actively antagonized the ability of IL-6 to rescue cells from that fate, consistent with its role as either a transcriptional repressor or activator, depending on the cell type in which it is expressed. We conclude that the E2A-HLF chimera abrogates p53-induced apoptosis in leukemic cells, possibly through the transcriptional modulation of cell death pathways that are activated by p53 in response to DNA damage.

Animals↗

PubMed1998

Mechanisms of elevation of serum and urinary concentrations of soluble thrombomodulin in diabetic patients: possible application as a marker for vascular endothelial injury.

Serum and urinary levels of soluble thrombomodulin (TM) were measured in 71 patients with non-insulin-dependent diabetes mellitus (NIDDM) and 132 age-matched control subjects to elucidate the mechanisms involved in increased TM levels. We compared the TM level with urinary albumin excretion (UAE), creatinine (Cr) clearance, and indices of renal tubular damage such as urinary beta2-microglobulin. Serum TM was significantly higher in diabetic patients versus control subjects (P < .001) regardless of whether the patients had diabetic nephropathy. Urinary TM levels were also higher in diabetic patients than in control subjects (P < .001). Serum TM in diabetic patients was correlated positively with serum Cr and UAE and inversely with the Cr clearance rate (P < .001, respectively). The urinary level of TM in diabetic patients was significantly correlated with 24-hour glucose excretion and the serum level of 1,5-anhydroglucitol (1,5-AG) (P < .001). However, no correlations were found between urinary TM levels and renal function in diabetic patients. There was also no correlation between serum and urinary levels of TM in the patients. These results suggest that although the serum TM level is influenced by an impairment of the renal clearance of TM, this parameter may be a useful marker for vascular endothelial injury in diabetic patients. On the other hand, since the elevated urinary level of TM in the patients paralleled their urinary excretion of glucose, urinary TM levels do not correlate with vascular endothelial injury in diabetic patients.

Adult↗

PubMed1998

Leukemic cells with 11q23 translocations express granulocyte colony-stimulating factor (G-CSF) receptor and their proliferation is stimulated with G-CSF.

We report a 20-month-old boy with acute lymphoblastic leukemia with the 11q23 translocation whose blasts markedly increased in peripheral blood after intravenous granulocyte colony-stimulating factor (G-CSF) administration, but disappeared after stopping G-CSF. The in vitro study showed that the leukemic cells separated from this patient expressed G-CSF receptor (G-CSFR) and an addition of G-CSF stimulated their proliferation by 3H-thymidine incorporation assay (stimulation index, 4.9). To clarify whether or not leukemic cells with 11q23 translocations generally express G-CSFR and show proliferative response to G-CSF, we performed the similar in vitro experiments using eight leukemic cell lines with 11q23 translocations. We found that all cell lines examined expressed G-CSFR (20-98%) and proliferation of seven leukemic cell lines was significantly enhanced in response to G-CSF (stimulation index >1.5 in five cell lines), suggesting a possible participation of the G-CSF/G-CSFR interaction in the process of growth regulation of leukemic cells with 11q23 translocations.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1998

Power spectral analysis of variations in heart rate in patients with hyperthyroidism or hypothyroidism.

Power spectral analysis (PSA) of the variation in heart rate is useful in determining the relative activity of the sympathetic and parasympathetic nerves. In this study, PSA was used to investigate the relationship between abnormalities in autonomic nerve function and the presence of thyroid disorders in patients with autoimmune thyroid diseases. The low frequency (LF) or high frequency (HF) components of R-R variations were determined by PSA. The coefficient of variation of the R-R time intervals (CV(R-R)) was positively correlated with HF in healthy subjects. In untreated hyperthyroid patients with Graves' disease, the CV(R-R) and HF values were significantly lower than in healthy controls. Moreover, the LF/HF ratio in patients with untreated Graves' disease was significantly higher, and the LF/HF ratio in hypothyroid patients with Hashimoto's thyroiditis was significantly lower than in healthy controls. A negative correlation was observed between serum levels of free thyroid hormones (FT4 and FT3) and HF in Graves' disease patients. In some hyperthyroid patients, antithyroid drug therapy or beta-blocker administration gradually restored reduced HF values. Present results suggest that relative vagal nerve activity is reduced in hyperthyroid patients and that this reduction is reversible according to the decrease in serum levels of thyroid hormones.

Adrenergic beta-Antagonists↗

PubMed1998

The AD1 and AD2 transactivation domains of E2A are essential for the antiapoptotic activity of the chimeric oncoprotein E2A-HLF.

The chimeric oncoprotein E2A-HLF, generated by the t(17;19) chromosomal translocation in pro-B-cell acute lymphoblastic leukemia, incorporates the transactivation domains of E2A and the basic leucine zipper (bZIP) DNA-binding and protein dimerization domain of HLF (hepatic leukemic factor). The ability of E2A-HLF to prolong the survival of interleukin-3 (IL-3)-dependent murine pro-B cells after IL-3 withdrawal suggests that it disrupts signaling pathways normally responsible for cell suicide, allowing the cells to accumulate as transformed lymphoblasts. To determine the structural motifs that contribute to this antiapoptotic effect, we constructed a panel of E2A-HLF mutants and programmed their expression in IL-3-dependent murine pro-B cells (FL5.12 line), using a zinc-inducible vector. Neither the E12 nor the E47 product of the E2A gene nor the wild-type HLF protein was able to protect the cells from apoptosis induced by IL-3 deprivation. Surprisingly, different combinations of disabling mutations within the HLF bZIP domain had little effect on the antiapoptotic property of the chimeric protein, so long as the amino-terminal portion of E2A remained intact. In the context of a bZIP domain defective in DNA binding, mutants retaining either of the two transactivation domains of E2A were able to extend cell survival after growth factor deprivation. Thus, the block of apoptosis imposed by E2A-HLF in pro-B lymphocytes depends critically on the transactivating regions of E2A. Since neither DNA binding nor protein dimerization through the bZIP domain of HLF is required for this effect, we propose mechanisms whereby protein-protein interactions with the amino-terminal region of E2A allow the chimera to act as a transcriptional cofactor to alter the expression of genes regulating the apoptotic machinery in pro-B cells.

Animals↗