A minimal glycine-alanine repeat prevents the interaction of ubiquitinated I kappaB alpha with the proteasome: a new mechanism for selective inhibition of proteolysis.

The Epstein-Barr virus nuclear antigen 1 contains a glycine-alanine repeat that inhibits in cis MHC class I-restricted presentation. We report here that insertion of a minimal glycine-alanine repeat motif in different positions of I kappaB alpha protects this NF-kappaB inhibitor from signal-induced degradation dependent on ubiquitin-proteasome, and decreases its basal turnover in vivo resulting in constitutive dominant-negative mutants. The chimeras are phosphorylated and ubiquitinated in response to tumor necrosis factor alpha, but are then released from NF-kappaB and fail to associate with the proteasome. This explains how functionally competent I kappaB alpha is protected from proteasomal disruption and identifies the glycine-alanine repeat as a new regulator of proteolysis.

Inactivation of the human fragile histidine triad gene at 3p14.2 in monochromosomal human/mouse microcell hybrid-derived severe combined immunodeficient mouse tumors.

We have previously shown that inoculation of human chromosome 3 (chr3)/A9 mouse fibrosarcoma microcell hybrids (MCHs) into severe combined immunodeficient (SCID) mice was followed by the regular elimination of some 3p regions whereas a 3q region was retained even after prolonged mouse passage. Using this approach, referred to as the elimination test (Et), we have defined a common eliminated region (CER) of approximately 7 cM at 3p21.3 that was absent in all of the 27 tumors generated from five MCHs. Later, CER was reduced to a 1-Mb region, designated as CER1. Another eliminated region (ER2) at 3p21.1-p14.2 was absent in 21 of the 27 tumors. ER2 borders at but does not include the fragile histidine triad (FHIT) gene, considered as a putative tumor suppressor gene. In the present work, two new and two previously studied MCHs, and 13 derived SCID mouse tumors were analyzed by fluorescence in situ hybridization (FISH) chromosome painting and by PCR, using 72 chr3p-specific and 11 chr3q-specific markers. Nine tumors generated from three MCHs that carried cytogenetically normal chr3, remained PCR-positive for all of the chr3 markers tested. Designated as "PCR+" tumors, they were examined by reverse transcription (RT)-PCR, together with four of six previously studied tumors derived from MCH910.7, which carried a del(3)(pter-p21.1), for the expression of 14 human genes: 5 genes within CER1 (LIMD1, CCR1, CCR2, CCR3, CCR5), 5 genes located within regions that were homozygously deleted in a variety of carcinomas (ITGA4L, LUCA1, PTPRG, FHIT, DUTT1), and 4 other genes in chr3p (VHL, MLH1, TGM4, UBE1L). We found that VHL, MLH1, ITGA4L, LIMD1, UBE1L, LUCA1, PTPRG, and DUTT1 were expressed in the MCH lines in vitro and also in the derived SCID tumors. No transcripts that originated from the four CCR genes or from TGM4 could be detected in any of the MCH lines. Alone among the 14 genes examined, FHIT showed a tumor growth-associated change. It was expressed in vitro in five of seven MCH lines. Nine of 13 derived tumors had no FHIT transcript. The remaining 4 expressed a truncated mRNA and a reduced amount of the full-length mRNA. We have previously found that FHIT was deleted at the DNA level in 17 of 21 tumors derived from four MCHs. The remaining 4 of 21 had no FHIT transcript. Our compiled data show that FHIT was either physically or functionally impaired in all 34 of the 34 analyzed tumors. Variants with deleted or down-regulated FHIT have a selective growth advantage.

The EBV-encoded LMP1 protein inhibits p53-triggered apoptosis but not growth arrest.

We have previously shown that exogenous wild type p53 induces apoptosis in the Burkitt lymphoma line BL41 that carries endogenous mutant p53, using a temperature sensitive p53 construct expressed as mutant p53 at 37 degrees C and wild type p53 at 32 degrees C (Ramqvist et al., Oncogene, 8, 1495-1500, 1993). We also found that wild type p53-induced apoptosis is blocked by bcl-2 in a mouse T lymphoma line (Wang et al., Oncogene, 8, 3427-3431, 1993) The Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) can protect Burkitt lymphoma cells from apoptosis induced by low serum. In order to test if LMP1 can block p53-triggered apoptosis, we infected BL41 cells expressing the ts p53 construct with an LMP1-carrying retrovirus. The LMP1-expressing BL41-ts p53 cells were arrested in G1 upon induction of wild type p53 expression at 32 degrees C, but did not enter apoptosis as shown by the absence of positive TUNEL staining. WAF1/p21 mRNA was induced at 32 degrees C in both the ts p53-expressing and ts p53/LMP1-expressing BL41 cells. Thus, LMP1 prevents p53-induced apoptosis but does not interfere with induction of WAF1/p21. The LMP1-infected cells expressed elevated bcl-2 protein levels. Therefore, our data suggest that LMP1 blocks p53-triggered apoptosis but not G1 arrest by upregulating bcl-2 expression.

Combined LOH/CGH analysis proves the existence of interstitial 3p deletions in renal cell carcinoma.

We have recently developed an allele titration assay (ATA) to assess the sensitivity and influence of normal cell admixture in loss of heterozygosity (LOH) studies based on CA-repeat. The assay showed that these studies are biased by the size-dependent differential sensitivity of allele detection. Based on these data, we have set up new criteria for evaluation of LOH. By combining these new rules with comparative genome hybridization (CGH) we have shown the presence of interstitial deletions in renal cell carcinoma (RCC) biopsies and cell lines. At least three out of 11 analysed RCC cell lines and three out of 37 biopsies contain interstitial deletions on chromosome 3. Our study suggests the presence of several regions on human chromosome 3 that might contribute to tumor development by their loss: (i) 3p25-p26, around the VHL gene (D3S1317); (ii) 3p21. 3-p22 (between D3S1260 and D3S1611); (iii) 3p21.2 (around D3S1235 and D3S1289); (iv) 3p13-p14 (around D3S1312 and D3S1285). For the first time, AP20 region (3p21.3-p22) was carefully tested for LOH in RCC. It was found that the AP20 region is the most frequently affected area. Our data also suggest that another tumor suppressor gene is located near the VHL gene in 3p25-p26.

Human wig-1, a p53 target gene that encodes a growth inhibitory zinc finger protein.

We previously identified a novel p53-induced mouse gene, wig-1, that encodes a 290 amino acid zinc finger protein (Varmeh-Ziaie et al., 1997). Here we have identified and characterized the human homolog of mouse wig-1. The human wig-1 protein is 87% identical to the mouse protein and contains three zinc finger domains and a putative nuclear localization signal. Human wig-1 mRNA and protein is induced following activation of wild type p53 expression in our BL41-ts p53 Burkitt lymphoma cells. Wig-1 is also induced in MCF7 cells following treatment with the DNA-damaging agent mitomycin C. Northern blotting detected low levels of wig-1 mRNA in normal human tissues. Fluorescence in situ hybridization mapped wig-1 to human chromosome 3q26.3-27. FLAG-tagged human wig-1 localizes to the nucleus. Ectopic overexpression of human wig-1 inhibits tumor cell growth in a colony formation assay. These results suggest that human wig-1 has a role in the p53-dependent growth regulatory pathway.

Spontaneous development of plasmacytomas in a selected subline of BALB/cJ mice.

Sixty per cent of BALB/cAnPt mice injected intraperitoneally (i.p.) with tetramethylpentadecane (pristane) develop plasmacytomas (PCs), whereas less than 10% of BALB/cJ develop such tumours. Most other mouse strains are completely resistant. Resistance is dominant over susceptibility in F1 hybrids between BALB/cAnPt and the resistant non-BALB/c strains, suggesting that susceptibility may be due to some genetic defect. (BALB/cAnPtxBALB/cJ)F1 hybrids have a PC incidence of 36-42%. Previously, BALB/cJ has been shown to harbour at least one resistance gene (Potter et al., Genomics 1988, Vol. 2, pp. 257-262). On the assumption that BALB/cJ may contain a segregating resistance gene, we cross BALB/cJ females with pristane-pretreated BALB/cJ males that were found to be carrying PC cells intraperitoneally 5-7 months after pristane treatment. After two selective crosses, 62% of the BALB/cJ subline BALB/cM2/22 developed PC after pristane and 52% after pristane followed by Abelson virus, while unselected controls had an incidence of 11% and 0%, respectively. Moreover, six spontaneous plasmacytomas developed in untreated females of the selected colony. Five of these carried T(12; 15) (F2; D2/3) translocations. The sixth had a T(1; 10) (G; C1) translocation and an interstitial duplication of segment (C1/E3) on one chromosome 5. It may be concluded that pristane treatment is not a prerequisite for the induction of the PC associated Ig/myc translocations.

Hypersomy of chromosome 15 with retrovirally rearranged c-myc, loss of germline c-myc and IgK/c-myc juxtaposition in a macrophage-monocytic tumour line.

From a lymphoid tumour induced by 7,12-dimethylbenz-[a]-anthracene (DMBA) + methyl-N-nitrose-N-urea (MNU) in an [AKR Rb(6.15) x CBAT6T6]F1 mouse, a macrophage- monocyte line (KT-10) was isolated. Following ethyl methanesulfonate (EMS) treatment, a bromodeoxyuridine (BUdR) resistant subline was selected. Serial propagation of this line in vitro in the presence of BUdR (28 months) with periodic cytogenetic and molecular examinations, has led to the definition of four successive stages. During stage I, the cells were trisomic for chromosome 15. They contained Rb(6.15) and Rb(del6.15) of AKR and T(14;15) of CBA origin. Southern blotting showed the presence of both germline (G) and rearranged (R) c-myc. At stage II, Rb(del6.15) has duplicated. Both Rb(6.15) and T(14;15) persisted together with G-myc and R-myc. In stage III, the CBA-derived T(14;15) was lost, in parallel with G-myc. At this stage, a Dic.In(6.15) was detected. One of its arms was cytogenetically identical with the long arm of In(6.15) in the variant IgK/myc translocations. This chromosome carried R-myc and IgK in juxtaposition, as indicated by comigration on pulsed field electrophoresis (PFGE). At stage IV, the R-myc carrying AKR-derived chromosome 15s were present in six copies. Possible relationships between the increasing R/G myc ratio and changed growth characteristics in vivo and in vitro are discussed.

Central and peripheral effects of 6-hydroxydopamine on exercise performance in rats.

6-Hydroxydopamine (6-OHDA) injected into the lateral cerebral ventricles of rats impaired their exercise performance until exhaustion (treadmill run and swimming). The injected rats displayed significantly less training-induced improvement of swimming performance than did the control animals. Reduced performance also was seen in adult rats treated intraperitoneally with 6-OHDA in the neonatal or adult periods. The functional impairment can be explained by the neurotoxic action of 6-OHDA on monoamine brain structures (substantia nigra and locus ceruleus) and/or on peripheral sympathetic innervation, the latter assessed by histofluorescence of the iris muscle. The possible contribution of lacticacidemia to the reduced tolerance to stress of 6-OHDA-treated rats by the intracerebroventricular route also is considered.

Sustained TGF beta exposure suppresses Smad and non-Smad signalling in mammary epithelial cells, leading to EMT and inhibition of growth arrest and apoptosis.

To better understand the dual, tumour-suppressive and tumour-promoting function of transforming growth factor-beta (TGFbeta), we analysed mammary epithelial NMuMG cells in response to short and long-term TGFbeta exposure. NMuMG cells became proliferation-arrested and apoptotic after exposure to TGFbeta for 2-5 days, whereas surviving cells underwent epithelial-mesenchymal transition (EMT). After chronic TGFbeta exposure (2-3 weeks), however, NMuMG cells became resistant to proliferation arrest and apoptosis, showing sustained EMT instead (TD cells). EMT was fully reversed by a pharmacologic TGFbeta-receptor-I kinase inhibitor or withdrawal of TGFbeta for 6-12 days. Interestingly, both cell cycle arresting/proapoptotic (Smads, p38 kinase) and antiapoptotic, proliferation and EMT-promoting signalling pathways (PI3K-PKB/Akt, ERK) were co-suppressed to low, but significant levels. Except for PI3K-Akt, TGFbeta-dependent downregulation of these signalling pathways in transdifferentiated (TD) cells was fully reversed upon TGFbeta withdrawal, together with partial re-induction of proliferation arrest and apoptosis. Co-injection of non-tumorigenic NMuMG cells with tumour-forming CHO cells oversecreting exogenous TGFbeta1 (CHO-TGFbeta1) allowed outgrowth of epithelioid cells in CHO-TGFbeta1 cell-induced tumours. These epithelial islands enhanced CHO-TGFbeta1 tumour cell proliferation, possibly due to chemokines (for example, JE/MCP-1) secreted by NMuMG/TD cells. We conclude that suppression of antiproliferative, proapoptotic TGFbeta signalling in TD cells may permit TGFbeta-dependent proliferation, survival and EMT-enhancing signalling pathways to act at low levels. Thus, TGFbeta may modulate its own signalling to facilitate switching from tumour suppression to tumour progression.

Evolutionarily plastic regions at human 3p21.3 coincide with tumor breakpoints identified by the "elimination test".

We have previously found with the microcell hybrid-based "elimination test" that human chromosome 3 transferred into murine or human tumor cells regularly lost certain 3p regions during tumor growth in SCID mice. The most common eliminated region, CER1, is approximately 2.4 Mb at 3p21.3. CER1 breakpoints were clustered in approximately 200-kb regions at both telomeric and centromeric borders. We have also shown, earlier, that tumor-related deletions often coincide with human/mouse synteny breakpoints on 3p12-p22. Here we describe the results of a comparative genomic analysis on the CER1 region in Caenorhabditis elegans, Drosophila melanogaster, Fugu rubripes, Gallus gallus, Mus musculus, Rattus norvegicus, and Canis familiaris. First, four independent synteny breaks were found within the CER1 telomeric breakpoint cluster region, comparing human, dog, and chicken genomes, and two independent synteny breaks within the CER1 centromeric breakpoint cluster region, comparing human, mouse, and chicken genomes, suggesting a nonrandom involvement of tumor breakpoint regions in chromosome evolution. Second, both CER1 breakpoint cluster regions show recent tandem duplications (seven Zn finger protein family genes at the telomeric and eight chemokine receptor genes at the centromeric side). Finally, all genes from these regions underwent horizontal evolution in mammals, with formation of new genes and expansion of gene families, which were displayed in the human genome as tandem gene duplications and pseudogene insertions. In contrast the CER1 middle region contained evolutionarily well-conserved solitary genes and a minimal amount of retroposed genes. The coincidence of evolutionary plasticity with CER1 breakpoints may suggest that regional structural instability is expressed in both evolutionary and cancer-associated chromosome rearrangements.

Visualization of alternative Epstein-Barr virus expression programs by fluorescent in situ hybridization at the cell level.

Epstein-Barr virus (EBV) transforms human B lymphocytes into immortalized lymphoblastoid cell lines (LCLs). They regularly express six virally encoded nuclear proteins (EBNA1 to EBNA6) and three membrane proteins (LMP1, LMP2A, and LMP2B). In contrast, EBV-carrying Burkitt lymphoma (BL) cells in vivo and derived type I cell lines that maintain the BL phenotype express only EBNA1. During prolonged in vitro culturing, most EBV-carrying BL lines drift toward a more immunoblastic (type II or III) phenotype. Their viral antigen expression is upregulated in parallel. We have used fluorescent in situ hybridization to visualize viral transcripts in type I and III BL lines and LCLs. In type I cells, EBNA1 is encoded by a monocistronic message that originates from the Qp promoter. In type III cells, the EBNA1 transcript is spliced from a giant polycistronic message that originates from one of several alternative Wp or Cp promoters and encodes all six EBNAs. We have obtained a "track" signal with a BamHI W DNA probe that could hybridize with the polycistronic but not with the monocistronic message in two type III BL lines (Namalwa-Cl8 and MUTU III) and three LCLs (LCL IB4-D, LCL-970402, and IARC-171). A BamHI K probe that can hybridize to both the monocistronic and the polycistronic message visualized the same pattern in the type III BLs and the LCLs as the BamHI W probe. A positive signal was obtained with the BamHI K but not the BamHI W probe in the type I BL lines MUTU I and Rael. The RNA track method can thus distinguish between cells that use a type III and those that use a type I program. The former cells hybridize with both the W and the K probes, but the latter cells hybridize with only the K probe. Our findings may open the way for studies of the important but still unanswered question of whether cells with type I latency arise from immunoblasts with a full type III program or are generated by a separate pathway during primary infection.

Nonrandom loss of human chromosome 3 fragments from mouse-human microcell hybrids following progressive growth in SCID mice.

Microcell hybrid lines of A9 mouse fibrosarcoma containing complete or partially deleted human chromosomes 3 (chr. 3) were inoculated into SCID mice. Cell lines derived from the tumors were examined by fluorescent in situ hybridization for the status of the transferred human chromosome and by PCR for marker loss. The SCID tumors arising after the inoculation of 10(5) cells were passaged serially in vivo and regularly showed loss of four markers; D3S1029 (3p21.3-21.2), AP20R (3p22-21.3, D3S32 (3p21.3-p21.2), and THRB (3p24). This regularly deleted region is bordered by markers GNA12 (3p21.1-p21.3) and VHL (3p25) that were maintained in a fraction of tumors. Fragments derived from the long arm of chromosome 3 and corresponding markers in the 3q26-q28 region were retained in all tumors. Our findings may be related to the postulated presence of tumor suppressor genes in the 3p24-p21 region as indicated by the frequent deletion of this region in renal and small cell lung carcinomas and other solid tumors. The technically cumbersome identification of suppressor genes may be supplemented by an "elimination test" based on analogous principles.

[Pathogenetic forms of intersexuality states in children]. / Forme patogenetice ale starilor de intersexualitate la copil.

The present paper reports on 26 children with intersexuality states, belonging to the following pathologic forms: 1. female pseudohermaphroditism--11 cases, including a) type I congenital corticoadrenal hyperplasia (8 cases) and type III (1 case); b) iatrogenic form (1 case), and c) corticoadrenal virilizing adenoma (1 case); 2. male pseudohermaphroditism--6 cases, and 3. gonadal dysgenesis of female phenotype--9 cases of which a) Turner syndrome (6 cases); b) gonadal dysgenesis 45 XO/46 XX (2 cases) and c) Swyer syndrome (1 case). The authors emphasize the prenatal conditioned character (chromosomal or metabolic genetic diseases, congenital diseases) in the majority of the cases; they discuss the diagnostic criteria, therapeutical possibilities and prophylaxis as well as their efficiency which depends upon the moment the diagnosis is established.

Correlation of myc expression with the growth-arrested and transformed phenotypes in hybrids between a T lymphoma and an antigen-responsive T-cell line.

Fusion of the YACUT lymphoma cell line with the Mls-1a-antigen-specific non-tumorigenic T-cell line G4 produced growth-arrested hybrids that could be induced to proliferate in the presence of Mls-1a antigen. Prolonged growth of such hybrids by repeated antigenic stimulation resulted in the appearance of autonomously growing hybrid lines. Of the 4 antigen-independent hybrid clones, I was weakly tumorigenic (25% incidence) while the other 3 were highly tumorigenic (100% incidence). In the growth-arrested hybrids the de-regulated c-myc expression characteristic of the YACUT cells was suppressed. In the autonomously growing clones, however, c-myc expression had reverted to the levels of the lymphoma parent and 1 to 2 extra copies of chromosome 15 were consistently present. These results indicate that repeated antigenic stimulation somehow abrogated the down-regulation of c-myc in the growth-arrested hybrid lines. The increase in the number of copies of chromosome 15, however, suggests that genes located on this chromosome may abolish the effect of the negative regulatory functions of the non-malignant parent in a gene-dosage-dependent manner.

Influence of thymic humoral factor(s) on haemopoiesis in mice.

Postirradiation administration of Leukotrophin to whole-body irradiated mice was associated with increased LD50/30 and DRF. As indicated by 59Fe uptake and ESC number, haemopoiesis was significantly stimulated in spleen and bone marrow after Leukotrophin application to irradiated mice. DNA content and the uptake of 3H-thymidine in DNA was significantly enhanced in the thymus and bone marrow of irradiated and Leukotrophin-treated mice. The micronucleus test confirmed that Leukotrophin is a therapeutic agents, while administered before irradiation it does not influence the initial radio-lesions.

New data concerning the effects of 6-hydroxydopamine on the exercise performance in rats.

An injection of 250 micrograms 6-OHDA into the right cerebral ventricle of adult rats tested after 45 days led to a reduction of the run performance on the treadmill compared with the control group, but only in the first period of this test, repeated 10-11 times, every 2-3 days. This behavior was more obvious in male and in exercise-enduring rats. The histological examination proved partial destruction of central monoaminergic (especially dopaminergic) structures in the 6-OHDA treated rats. 24-48 hours after peripheral injections of 6-OHDA (100 mg/kg b. w.) a considerable reduction of the run and swim performance (acute exercise till exhaustion) occurred. Falck's histofluorescence method proved large destructions of the noradrenergic innervation in the iris; 6-OHDA injected i.p. into neonatal rats had similar effects.

B cell phenotype-dependent expression of the Epstein-Barr virus nuclear antigens EBNA-2 to EBNA-6: studies with somatic cell hybrids.

The expression of the transformation-associated Epstein-Barr virus (EBV)-encoded nuclear antigens (EBNAs) 1 to 6 and membrane protein LMP-1 was studied in a series of somatic cell hybrids derived from the fusion of the EBV-transformed lymphoblastoid cell line (LCL) KR-4, and the EBV-carrying Burkitt's lymphoma lines Daudi, P3HR-1 and Raji, with non-B cell lines of fibroblast, erythroid, myeloid and epithelial origin. Expression of EBNAs 2 to 6 was down-regulated in the hybrids in parallel with extinction of the B cell markers CD19, CD20, CD21, CD23, HLA class II, and surface or cytoplasmic immunoglobulin. LMP-1 was expressed independently of EBNA-2 in hybrids derived by the fusion of the LMP-1-positive KR-4 and P3HR-1 cell lines with epithelial and myeloid cells, respectively. LMP-1 was down-regulated in hybrids derived by the fusion of P3HR-1 with an erythroid cell line and in the hybrid between Raji and a mouse fibrosarcoma line. EBNA-1 was the only EBV antigen that was regularly expressed in the hybrids regardless of the dominating cellular phenotype. The autonomous expression of EBNA-1 suggests that its regulatory pathway is independent of phenotype-associated cellular or viral factors. In contrast, the expression of EBNAs 2 to 6 appears to require a B cell environment. EBNA-2 was shown to contribute to the regulation of LMP expression in B cells. We show that in LCL-carcinoma hybrids the dominating epithelial phenotype is permissive for LMP expression in the absence of EBNA-2.

The LZTFL1 gene is a part of a transcriptional map covering 250 kb within the common eliminated region 1 (C3CER1) in 3p21.3.

Deletions on 3p have been described in a large number of human tumors, suggesting the presence of a tumor suppressor gene(s). Using the elimination test, we previously defined a 1-Mb segment from human 3p21.3 (C3CER1). Genomic sequencing allowed us to construct a transcription map covering 250 kb containing five genes. We have characterized a human leucine zipper containing gene, leucine zipper transcription factor-like 1 (LZTFL1), and its mouse orthologue (Lztfl1), which was also mapped to mouse chromosome 9F. The LZTFL1 gene has two transcript isoforms displaying alternative polyadenylation. We have localized the human orthologue of the yeast SAC1 (suppressor of actin) gene as well as characterized and mapped the mouse Sac1 gene. Furthermore, the XT3 gene was characterized, encoding a member of the Na(+)/Cl(-) neurotransmitter superfamily. It has been shown that the XT3 gene had an alternatively spliced brain-specific isoform, predicted to remove 1 of 12 putative transmembrane domains. The transcription map also includes the CC chemokine receptor 9 gene (CCR9) and the LIM domain containing gene 1 (LIMD1). This work partially defines the gene content of C3CER1 that is a prerequisite for delineation of its role in tumorigenesis.

Restoration of H-2b expression and processing of endogenous antigens in the MHC class I pathway by fusion of a lymphoma mutant to L cells of the H-2k haplotype.

The RMA-S lymphoma mutant cannot process and present antigens to H-2-restricted cytotoxic T lymphocytes. It synthesizes major histocompatibility complex class I heavy (H-2KbDb) and light beta 2-microglobulin (beta 2mb) chains of normal size and charge, but only a fraction of these assemble and reach the cell surface. As a first step investigating the genetic defect of this line, we have fused it to a L cell fibroblast line (H-2KkDk/beta 2ma). The fusion restored H-2Kb, Db and beta 2mb expression as well as the ability to process and present internally derived (minor histocompatibility and influenza virus nucleoprotein) antigens in RMA-S. This shows that the mutation(s) responsible for the phenotype of RMA-S is (are) not located within the MHC class I heavy and light chain genes. Other cellular factors, derived from the L cell fusion partner, can control antigen processing and transport of MHC class I molecules. These findings are discussed in relation to the observation that assembly and transport of MHC class I molecules can be induced in the mutant by H-2b-restricted peptides. The recessive nature of the defect and its independence of MHC class I genes in the mutant has important implications for future transfection studies, of this and similar mutants, aiming at establishing cells containing non-assembled MHC class I molecules of different alleles and identifying the gene(s) controlling processing of endogenous antigens.

Involvement of 3p deletions in sporadic and hereditary forms of renal cell carcinoma.

Deletions of the short arm of chromosome 3 and associated allele losses have been reported in the majority of sporadic renal cell carcinomas (RCC). On the basis of the combined cytogenetic and molecular data, it is reasonable to assume that a putative RCC locus, which contributes to tumor development by its loss, is located telomerically of the D3F15S2 site. Using H3E4, a D3F15S2-specific probe, we have isolated a cDNA clone (cl.4-2), and a sequence comparison revealed that the cDNA clone corresponds to the human acyl-peptide hydrolase gene. The gene is fairly universally expressed, but in RCC biopsies its expression is severely reduced, compared to the normal kidney. Cl.4-2 was used for in situ hybridization on metaphase chromosomes prepared from an Epstein-Barr virus (EBV) transformed lymphoblastoid cell line, derived from a t(3;8) (p14.2;q24.1) carrying member of the RCC family described by Cohen et al. in 1979 (N Engl J Med: 301:592-595). Carriers of this translocation regularly develop RCC by middle age. We now report that D3F15S2 is localized on the telomeric side of the constitutional breakpoint, in 3p21. The region of 3p affected by this familial translocation is thus not identical with the region of 3p most frequently deleted in sporadic RCC.