Chromosome Assignment of the T-Antigen Gene of Simian Virus 40 in African Green Monkey Cells Transformed by Adeno 7-SV40 Hybrid

Somatic cell hybrids between mouse cells deficient in thymidine kinase [ATP:thymidine 5′-phosphotransferase (EC 2.7.1.75)] and two different monkey cell lines transformed by an adeno 7-SV40 hybrid have been produced using both a semiselective and a double selective procedure. Concordant segregation of the expression of SV40 T antigen with a specific monkey chromosome has been observed in all the mouse-monkey hybrid clones examined. Subcloning of three SV40 T antigen positive hybrid clones resulted in their segregation into SV40 T antigen-positive and negative subclones. Positive correlation between the SV40 T antigen and the same monkey chromosome has been observed in all the subclones examined.     

Assignment of gene(s) for cell transformation to human chromosome 7 carrying the simian virus 40 genome.

Somatic cell hybrid clones between either C57BL/6 or Balb/c mouse peritoneal macrophages and two different simian virus 40 (SV40)-transformed human cell lines deficient in hypoxanthine phosphoribosyltransferase (EC 2.4.2.8; IMP:pyrophosphate phosphoribosyltransferase) were obtained in hypoxanthine-aminopterin-thymidine selective medium. All the hybrid cell clones contained the human chromosome 7, which carries the SV40 genome, and were SV40 tumor (T)-antigen positive. No hybrid cell clones studied displayed the density-dependent inhibition of cell growth characteristic of normal cells; all clones had a high saturation density and gave origin to cell colonies when plated in soft agar. Since the expression of the transformed phenotype was always associated with the presence of the human chromosome 7, which carries the SV40 genome, it is concluded that this chromosome contains gene(s) [Tr gene(s)] coding for "transforming factor(s)."     

Assignment of the integration site for simian virus 40 to chromosome 17 in GM54VA, a human cell line transformed by simian virus 40.

GM54VA human cells transformed by simian virus 40 (SV40) were fused with peritoneal macrophages obtained from three different mouse strains. All 27 hybrid clones studied were positive for SV40 tumor antigen in 100% of their cells and contained human chromosome 17. Human chromosome 17 was the only human chromosome present in five of the hybrid clones. Fusion of GM54VA cells and either thymidine kinase (EC 2.7.1.75)-deficient mouse or Chinese hamster fibroblasts resulted in the growth in hypoxanthine-aminopterin-thymidine medium of hybrid clones positive and negative for SV40 tumor antigen. Counterselection of the hybrid clones positive for tumor antigen in medium containing 5-bromodeoxyuridine resulted in the growth of hybrid cells that were negative for tumor antigen. These experiments indicate that negative for tumor antigen. These experiments indicate that SV40 is integrated in only one of the two parental human chromosomes 17. Because the genome of SV40 has been assigned to human chromosome 7 in two other SV40-transformed human cell lines, at least two different integration sites for SV40 would seem to be present in human cells: one located in human chromosome 7 and the other located in human chromosome 17.     

Somatic cell hybrids producing antibodies specific for the tumor antigen of simian virus 40.

We have produced somatic cell hybrids between mouse myeloma cells deficient in hypoxanthine phosphoribosyltransferase IMP: pyrophosphate phosphoribosyltransferase; EC 2.4.2.8) and spleen cells derived from mice primed with either syngeneic or allogeneic cells transformed by simian virus 40. Such hybrids produced antibodies specific for simian virus 40 tumor (T) antigen. Only four of twelve independent hybrid cell cultures produced antibodies against simian virus 40 T antigen that crossreacted with the T antigen induced by BK virus, a human papovavirus isolated from patients who had undergone immunosuppressive therapy.     

Genetics of type II glycogenosis: Assignment of the human gene for acid α-glucosidase to chromosome 17

We have studied somatic cell hybrids between thymidine kinase (EC 2.7.1.75) deficient mouse cells and human diploid fibroblasts for the expression of human acid alpha-glucosidase (EC 3.2.1.20). A deficiency in this enzyme is associated with the type II glycogenosis or Pompe disease. All 30 somatic cell hybrids selected in hypoxanthine/aminopterin/thymidine medium expressed human acid alpha-glucosidase and galactokinase (EC 2.7.1.6) and retained human chromosome 17; counterselection of the same hybrids in medium containing 5-bromodeoxyuridine resulted in the growth of hybrids that concordantly lost the expression of human acid alpha-glucosidase and galactokinase as well as human chromosome 17. Hybrids between thymidine kinase-deficient mouse cells and fibroblasts from a patient with Pompe disease that contained human chromosome 17 were found not to express human acid alpha-glucosidase. Because we have already shown that hybrids between mouse peritoneal macrophages and GM54VA simian virus 40-transformed human cells selectively retain human chromosome 17 and lose all other human chromosomes, we tested 13 independent mouse macrophage x GM54VA hybrid clones, including two that retained human chromosome 17 and no other human chromosomes, for the expression of human acid alpha-glucosidase and galactokinase. All 13 hybrid clones were found to express these human enzymes. Thus, we conclude that the gene coding for human acid alpha-glucosidase is located on human chromosome 17.     

Another chromosomal assignment for a simian virus 40 integration site in human cells.

Somatic cell hybrids derived from fusion of GM637, a human cell line transformed by simian virus 40, and mouse B82 cells were examined for simian virus 40 T antigen, V antigen, and viral DNA. All hybrid cell lines that contained viral DNA were T-antigen positive. Cells that did not have viral DNA were T-antigen negative. We determined that there is a single viral insertion in these hybrid cells. Correlation of T-antigen expression and viral DNA with the partial complements of the human genome retained in the hybrids shwed that the inserted viral genome is in human chromosome 8. The integrated viral DNA is stable; free viral DNA found in GM637 does not insert at other potential sites in the human genome.     

Somatic cell hybrids between mouse peritoneal macrophages and simian-virus-40-transformed human cells: II. Presence of human chromosome 7 carrying simin virus 40 genome in cells of tumors induced by hybrid cells.

Cells derived from tumors induced in "nude" mice after injection of cells that were hybrids between mouse peritoneal macrophages and simian virus 40 (SV40)-transformed human cells were found to retain the human chromosome 7 carrying the SV40 genome, and indicate that the presence of human chromosome 7 carrying the SV40 genome is responsible for the expression of the tumorigenic phenotype in the hybrid cells.     

Simian virus 40-related antigens in three human meningiomas with defined chromosome loss.

Two out of seven meningiomas tested in early cell cultures by indirect immunofluorescence staining showed simian virus 40 (SV40)-related tumor (T) antigen. In one tumor 90% of the cells were positive. An additional SV40-related antigen (U) was found in 10% of cells of a third tumor. These findings indicate that the meningioma cells showing a positive reaction are transformed by a papova virus that has at least partly the same antigenic properties as SV40 virus. SV40-related viral capsid (V) antigen was absent in all the meningiomas tested. No virus infectious for African green monkey kidney (AGMK) cells could be isolated. The tumors positive for T and U antigens showed the chromosome aberration typical for human meningiomas, i.e., the loss of one chromosome, G-22. The T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive meningioma cells were performed: fusion of cells pretreated with 8-azaguanine with cells premissive for SV40 led to a low percentage (0.01-0.05%) of V-antigen-positive nuclei in heterokaryon cultures. On the basis of these results, the possibility of a correlation between the meningioma, a relatively common intracranial tumor in man, and an SV40-related papova virus must be considered. It remains to be shown whether this virus is a causative agent for human meningiomas.     

Expression of human adenosine deaminase after fusion of adenosine deaminase-deficient cells with mouse fibroblasts

Two human choriocarcinoma cell lines were shown to be deficient in adenosine deaminase (ADA; adenosine aminohydrolase, EC 3.5.4.4) such that they did not produce bands on starch gels after electrophoresis and histochemical staining. Radiometric assay indicated that their ADA specific activity was approximately 2% that of HeLa (human) cell controls. Subclone analysis of one of the lines indicated that this deficiency was representative of individual cells of the line. After fusion of these cells with mouse fibroblasts having high ADA activity, most independently isolated hybrid clones expressed one of two, or both, additional (to the mouse) bands of ADA activity after electrophoresis. The expression of these extra bands in hybrids was dependent upon actual fusion. The phenomenon was observed in 30 of 45 independently derived hybrid clones from four different fusion experiments involving two different parental lines from each species. The pattern of appearance of the extra bands in independent hybrid clones and the tendency of a hybrid clone to lose one of the extra bands through subsequent passages suggests that the bands were the products of human genetic material. The extra bands electrophoretically comigrated with human ADA 1 and 2 from human ADA-1-2 heterozygotes and the faster-migrating of the two extra bands comigrated with human ADA 1 from HeLa cells. Therefore, we suggest that the bands appearing in hybrids are the products of the 1 and 2 alleles of the human ADA locus. The human cells used for fusion were deficient in ADA activity but contained the genetic information for ADA 1 and 2. Fusion with mouse cells having ADA activity resulted in the activation of both human gene products coded for on separate homologous chromosomes. We conclude that the human ADA locus is under manipulatable genetic regulation.     

Cytological Mapping of Human X-Linked Genes by Use of Somatic Cell Hybrids Involving an X-Autosome Translocation

Man-mouse and man-Syrian hamster somatic hybrid cell lines were prepared by fusion of mouse A9 or hamster TG2 cells, which are deficient in hypoxanthine-guanine phosphoribosyl transferase, with cells of a diploid fibroblastic strain, KOP-1, derived from a woman heterozygous for an X-autosome translocation. 61 clones were derived in nonselective medium and 85 sublines of these were derived in selective media: 53 in hypoxanthine-aminopterine-thymidine and 32 in 8-azaguanine. All three human X-linked markers studied, i.e., hypoxanthineguanine phosphoribosyl transferase (EC 2.4.2.8), glucose-6-phosphate dehydrogenase (EC 1.1.1.49), and phosphoglycerate kinase (EC 2.7.2.3), were present together, or absent together, in most of these clones and sublines. However, loss or retention of only phosphoglycerate kinase was occasionally observed, even in the absence of selective growth, while no evidence of separation of hypoxanthine-guanine phosphoribosyl transferase from glucose-6-phosphate dehydrogenase occurred. Cytological examination of eight man-hamster clonal lines by the quinacrine fluorescent technique showed that human phosphoglycerate kinase was only present when the translocation chromosome carrying most of the long arm of the X chromosome was present. The presence of human glucose-6-phosphate dehydrogenase and hypoxanthine-guanine phosphoribosyl transferase was not related to the presence or absence of this chromosome, but appeared to be correlated with the presence of the other translocation chromosome.     

Regional gene assignment of human porphobilinogen deaminase and esterase A4 to chromosome 11q23 leads to 11qter.

The regional gene assignments for human porphobilinogen deaminase (PBGD; EC 4.3.1.8) and esterase A4 (ESA4; EC3.1.1.1) chromosome 11 have been determined with somatic cell hybridization and immunologic, electrophoretic, and cytogenetic techniques. Dimethyl sulfoxide-induced erythroid differentiation of hybrid clones derived from the fusion of tetraploid Friend murine erythroleukemia (2S MEL) cells deficient in thymidine kinase and human Lesch--Nyhan fibroblasts (HLN) deficient in hypoxanthine phosphoribosyltransferase (HPRT-; EC 2.4.2.8) were examined for expression of human PBGD, ESA4, and lactate dehydrogenase A (LDHA; EC 1.1.1.27). Human PBGD was detected by rocket immunoelectrophoresis with rabbit anti-human PBGD IgG and by isoelectric focusing. The human chromosome complement of each clone was determined by cytogenetic and enzyme marker analyses. Of the five primary 2S MEL--HLN clones examined, three were positive for human PBGD. These were subcloned to yield a total of 10 secondary, tertiary, or quaternary clones. Analyses of these subclones permitted the regional assignment of human PBGD and ESA4 to the long arm of chromosome 11. Finer regional assignment of the loci for human PBGD and ESA4 was facilitated when two 2S MEL (HPRT-)--human fibroblast (HX/11) hybrids, each containing the X chromosome--autosome translocation (der11), t(X;11)(q25-26;q23) as the only human chromosome, were examined for expression of human PBGD, ESA4, and LDHA. One clone, HX/11-2, contained the intact X/11 translocated chromosome; in the other, HX/11-3, 11p was deleted, and the human X/11 derivative was translocated onto a mouse chromosome. HX/11-2 expressed human LDHA, but HX/11-3 did not, verifying that the latter human 11/X derivative did not include 11pter leads to 11p12; PBGD and ESA4 were not detected in either hybrid. These results confirm the location of the gene for human PBGD on chromosome 11 and establish the assignment of the loci for PBGD and ESA4 in the region 11q23 leads to 11qter.     

Chimeric mice derived from human—mouse hybrid cells

Mouse teratocarcinoma cells from the OTT6050 ascites tumor were established in tissue culture and selected for 5-bromodeoxyuridine (BrdUrd) resistance. The embryonal carcinoma cells grew without a feeder layer, remained deficient for thymidine kinase (EC 2.7.1.75), and differentiated like the original tumor into various tissues after subcutaneous injection into 129 mice. We fused the BrdUrd-resistant mouse teratocarcinoma cells with HT1080-6TG human diploid fibrosarcoma cells deficient in hypoxanthine phosphoribosyltransferase (EC 2.4.2.8) and selected for hybrid cells in hypoxanthine/aminopterin/thymidine medium. The resulting hybrid cells segregated human chromosomes quickly and retained one to three human chromosomes including chromosome 17 that carries the human genes for thymidine kinase and galactokinase (EC 2.7.1.6). Single hybrid cells from five independent clones containing human chromosome 17 were injected into mouse blastocysts bearing several genetic markers that affect the coat color phenotype and strain-specific enzyme variants in order to detect tissue differentiation derived from the injected cells. After the injection of single hybrid cells into a total of 103 experimental blastocysts that had been surgically transferred to pseudopregnant foster mothers, 49 mice were born and 2 of them clearly revealed coat mosaicism. In 2 of 17 mice thus far analyzed, the injected hybrid cells proved to be capable of participating substantially in development of seven different organs. However, human gene products have not yet been detected unequivocally in those tissues and weak human-specific galactokinase activity could be recovered only from two mosaic tissues.Our results demonstrate that, after in vitro culture and selection, at least some of the human-mouse hybrid cells still retain their in vivo potential to differentiate and become functionally integrated in the living organism. It now seems feasible to cycle mouse teratocarcinoma cells carrying human genetic material through mice via blastocyst injection to study human gene expression during differentiation.     

Expression of human myeloid-associated surface antigens in human-mouse myeloid cell hybrids.

Hybrid cell lines were obtained after fusion of mouse myeloid cells (WEHI-TG) with leukocytes from two patients with chronic myeloid leukemia. A third fusion was carried out with leukocytes from a patient with acute lymphocytic leukemia. All three patients carried the Philadelphia chromosome (Ph1) in the leukemia cell population. Cytochemical analysis confirmed the myelo-monocytic nature of the hybrid cell lines. The presence of Ph1 translocation products could be established in most hybrids derived from the two chronic myeloid leukemic patients, which confirms that indeed human myeloid cells were fused. Several of these hybrid lines showed reactivity with monoclonal antibodies known to be specific for human myeloid cells, whereas interlineage Chinese hamster fibroblast-human chronic myeloid leukemia hybrids failed to react with these antibodies. Five independently obtained monoclonal antibodies--MI/NI, UJ-308, VIM-D5, FMC-10, and B4.3--showed very similar reactivity patterns when tested on the hybrid clones. This result substantiates the evidence obtained from other studies, that these five antibodies are directed against the same myeloid-associated antigen. The gene(s) for expression of the latter antigen could be assigned to human chromosome 11.     

Cytoplasmic transfer of DNA containing simian virus 40 sequences into mouse 3T3 cells.

This paper describes the rare cytoplasmic transmission of defective simian virus 40 (SV40) viral DNA from enucleated cells (i.e., cytoplasts) of the SV40-transformed mouse cell line SVT2 (chloramphenicol-resistant) into cybrid cells formed by fusion of these cytoplasts with BALB/c 3T3 cells (thymidine kinase-deficient). The cybrids were selected in medium containing 1% serum, bromodeoxyuridine, and chloramphenicol. They were identified by their 3T3 chromosome content, by the instability of tumor (T)-antigen expression, by their transformed phenotype, and by their drug resistance. The yield of rare cybrids was about 5 x 10(-7) 0.1% of the yield on medium with 10% serum. The presence of the SV40 genome was detected by the expression of SV40-specific T antigen and confirmed (unpublished data) by hybridization of viral DNA probes with restriction enzyme fragments of nuclear DNAs from cybrid clones. Restriction site mapping (unpublished data) showed that at least 1 kilobase of host flanking DNA on each side of the SV40 DNA was included in the transferred segment. The transforming DNA was not stably integrated initially, as judged by cellular heterogeneity in T-antigen expression. Stable T-antigen-positive and negative subclones were recovered in 10% serum; instability could be retained for at least 30 doublings during growth in 1% serum. The instability is interpreted as evidence of non-integration or unstable integration of the transferred DNA into the host genome. The cytoplasmic transfer is interpreted as evidence that chromosomal fragments or intact chromosomes can be transferred rarely through the cytoplasm in cybrid crosses.     

Assignment of a locus required for flavoprotein-linked monooxygenase expression to human chromosome 2.

Hybrid clones segregating human chromosomes were prepared by fusing mouse RAG cells to fresh human bone marrow cells and tested for the mixed-function oxygenase [flavoprotein-linked monooxygenase; RH, reduced-flavoprotein:oxygen oxidoreductase (RH-hydroxylating); EC 1.14.14.1] arylhydrocarbon hydrocarbon hydroxylase. Neither constitutive nor induced aryl hydrocarbon hydroxylase activity was detected in parental RAG cells. Induced aryl hydrocarbon hydroxylase was expressed in 4 out of 12 primary and 12 out of 19 secondary hybrid clones examined. Constitutive hydroxylase activity was detectable in 9 of the 15 inducible clones. All of the hybrid clones that exhibited constitutive hydroxylase activity were also inducible. There was a positive correlation between constitutive and induced hydroxylase activities although the absolute levels of the enzyme showed a wide range between different clones. Isozyme analysis performed on 12 primary and 19 secondary hybrid clones showed that aryl hydrocarbon hydroxylase activity was concordant with the expression of the human isozymes malate dehydrogenase (EC 1.1.1.37) and isocitrate dehydrogenase (EC 1.1.1.42), previously assigned to human chromosome 2. Isozyme markers for 19 other human chromosomes segregated independently from aryl hydrocarbon hydroxylase activity. The results suggest that the gene(s) required for aryl hydrocarbon hydroxylase activity are located on human chromosome 2.     

Sialidosis and galactosialidosis: chromosomal assignment of two genes associated with neuraminidase-deficiency disorders.

The inherited human disorders sialidosis and galactosialidosis are the result of deficiencies of glycoprotein-specific alpha-neuraminidase (acylneuraminyl hydrolase, EC 3.2.1.18; sialidase) activity. Two genes were determined to be necessary for expression of neuraminidase by using human-mouse somatic cell hybrids segregating human chromosomes. A panel of mouse RAG-human hybrid cells demonstrated a single-gene requirement for human neuraminidase and allowed assignment of this gene to the (pter----q23) region of chromosome 10. A second panel of mouse thymidine kinase (TK)-deficient LM/TK- -human hybrid cells demonstrated that human neuraminidase activity required both chromosomes 10 and 20 to be present. Analysis of human neuraminidase expression in interspecific hybrid cells or polykaryocytes formed from fusion of mouse RAG (hypoxanthine/guanine phosphoribosyltransferase deficient) or LM/TK- cell lines with human sialidosis or galactosialidosis fibroblasts indicated that the RAG cell line complemented the galactosialidosis defect, but the LM/TK- cell line did not. This eliminates the requirement for this gene in RAG-human hybrid cells and explains the different chromosome requirements of these two hybrid panels. Fusion of LM/TK- cell hybrids lacking chromosome 10 or 20 (phenotype 10+,20- and 10-,20+) and neuraminidase-deficient fibroblasts confirmed by complementation analysis that the sialidosis disorder results from a mutation on chromosome 10, presumably encoding the neuraminidase structural gene. Galactosialidosis is caused by a mutation in a second gene required for neuraminidase expression located on chromosome 20.     

Electron-Nuclear Double Resonance of a Protein That Contains Copper: Evidence for Nitrogen Coordination to Cu(II) in Stellacyanin

The study of hybrids from three crosses between mouse cells and SV40-transformed human cells have established a positive correlation between the loss of human chromosomes and that of the SV40-induced T-antigen from the hybrid cells. These results, as well as those of other workers, provide strong support for the hypothesis of the integration of the SV40 genome in the chromosomes of transformed cells. Further, it has been shown that hybrid cells which have lost T-antigen are capable of synthesizing this antigen upon infection with SV40, thereby demonstrating that loss of the viral antigen from the hybrid cells is not due to loss of some cellular gene required for the expression of the viral genome. Results of karyological analyses of the hybrid cells argue against the existence of a single specific integration site for the SV40 genome in human cells.     

Localized Derepression on the Human Inactive X Chromosone in Mouse-Human Cell Hybrids.

Evidence for derepression of the gene for hypoxanthine phosphoribosyltransferase (HPRT; IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8) on the human inactive X chromosome was obtained in hybrids of mouse and human cells. The mouse cells lacked HPRT and were also deficient in adenine phosphoribosyltransferase (APRT; AMP: pyrophosphate phosphoribosyltransferase; EC2.4.2.7). The human female fibroblasts were HPRT-deficient as a consequence of a mutation on the active X but contained a normal HPRT gene on the inactive X. The two human X chromosomes were further distinguished by differences in morphology: the inactive X was morphologically normal while the active X included most of the long arm of autosome no. 1 translocated to the distal end of the X long arm. Forty-one hybrid clones were first isolated by selection for the presence of APRT; when these clones were selected for HPRT, six of them yielded derivatives having human HPRT with incidences of about 1 in 10-6 APRT-selected hybrid cells. The HPRT-positive derivatives contained a normal-appearing X chromosome indistinguishable from the inactive X of the parental human fibroblasts. The active X with the translocation was not found in any of the HPRT-positive hybrid cells. Human phosphoglycerokinase (ATP:3-phospho-D-glycerate 1-phosphotransferase. EC 2.7.2.3) and glucose-6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP 1-oxidoreductase, EC 1.1.1.49), which are specified by X-chromosomal loci, were not detected in the hybrids expressing HPRT even though they contained an apparently intact X chromosome. The observations are most simply explained by the infrequent, stable derepression of inactive X chromosome segments that include the HPRT locus but not the phosphoglycerokinase and glucose-6-phosphate dehydrogenase loci.     

Epstein-Barr virus and human chromosomes: close association of the resident viral genome and the expression of the virus-determined nuclear antigen (EBNA) with the presence of chromosome 14 in human-mouse hybrid cells.

Fourteen hybrid clones derived from the fused cultures of human lymphoblastoid FV5 cells and 5-bromodeoxyuridine-resistant mouse fibroblastic MCB2 cells grown in hypoxanthine/aminopterin/thymidine selective medium were examined for the presence of Epstein-Barr virus (EBV) DNA, the expression of the virus-determined nuclear antigen (EBNA), and the presence of human chromosomes, in the course of serial passage in vitro. Among the hybrid clones tested, 3 were positive for EBV DNA and EBNA, whereas the remaining 11 were totally negative. The chromosome investigations showed that human chromosome 14 was consistently involved in all three EBV genome-positive and EBNA-positive hybrid clones, but not in any negative clones. In 10 subclones isolated from 1 of the 3 positive clones, all of which contained only chromosome 14 of the human chromosomes, a concordant segregation of EBNA, EBA DNA, and chromosome 14 was evident. These findings suggest that the resident EBV genome is closely associated with chromosome 14 and the presence of this particular chromosome alone is sufficient for the maintenance and the expression of EBV genetic information in human lymphoblastoid cells.     

Somatic cell genetic analysis of human cell surface antigens: chromosomal assignments and regulation of expression in rodent-human hybrid cells.

The expression of 13 newly defined human cell surface antigens identified by monoclonal antibodies was studied in a panel of reduced rodent-human somatic cell hybrid clones. For each antigenic system the segregation of antibody reactivity was concordant with the segregation of a specific human chromosome, permitting the chromosomal assignment of 13 gene loci determining antigen expression. The antigens can be placed in four groups on the basis of their patterns of control in the hybrid cells. (i) Presence of a single human chromosome is necessary and sufficient for antigen expression; L230 (assigned to chromosome 2), AJ425, K15 (chromosome 3), SR84 (chromosome 5), JF23, Q14 (chromosome 11), SV13 (chromosome 15), and F10 (chromosome 19). (ii) AJ2 (chromosome 10) and J143 (chromosome 17); two antigens coded for by separate human chromosomes but associated as a molecular complex on the surface of AJ2+/J143+ human cells. (iii) F8 (chromosome 19); antigen expression dependent on the growth characteristics of hybrid cells: substrate-adherent cells are F8+, whereas cells growing in suspension are F8-. (iv) AO122 and F23 (chromosome 15); antigen expression controlled by the permissive/inducing vs. nonpermissive/noninducing nature of the rodent fusion partner. Hybrids derived from both antigen-positive and antigen-negative human cells can express AO122 and F23 but only when specific rodent cell types are used for hybridization: N4TG-1 neuroblastoma and L cells, but not RAG renal carcinoma cells, permit AO122 expression, whereas RAG and L cells, but not N4TG-1 cells, permit F23 expression. The rapidly expanding list of monoclonal antibodies defining human cell surface molecules provides a range of markers to probe the genetic regulation of antigen diversity in somatic cells.