Hydrophobic Transmembrane Region Research Articles

Characterization of monoclonal antibodies to human immunodefiency virus type 1 gp41 by HIV-1 polypeptides expressed inEscherichia coli

Two monoclonal antibodies (MAbs) were produced in Balb/c mice by immunization with recombinant gp41 derived from expression of lambda-BH10 cDNA of the human immunodeficiency virus-1 (HIV-1) in the prokaryotic expression vector pEX-41. Characterization of the epitopes recognized by these MAbs was done with HIV-1 envelope (env) fusion proteins expressed in Escherichia coli encoding ten distinct segments of the env proteins. In comparison, another mouse MAb, M25, a human MAb directed against gp41, which was produced by the xeno hybridoma line 3D6 and a pool of human patient sera containing antibodies to HIV-1 were tested. We were able to demonstrate that the epitopes recognized by our MAbs are located between arg732 and ser759 of the HIV-1 env glycoprotein gp160 of HTLV-III strain B. M25 reacted with epitopes between ser647 and pro731, which includes the hydrophobic transmembrane region of gp41. The human MAb against gp41, 3D6 reacts with epitopes between ile474 and trp646, a polypeptide stretch consisting of gp120 and gp41 specific amino acids. The human serum pool, positive for HIV-1 antibodies, reacted predominantly with antigenic determinants located between ile474 and leu863. The recombinant env fusion proteins were initially produced to test the immunoreactivity with patient sera and to characterize epitopes which are relevant for immunodiagnostic purposes. In this study, we showed that the set of recombinant env proteins is also a simple and accurate tool for the characterization of MAbs directed to the HIV envelope proteins.

Characterization of monoclonal antibodies to human immunodefiency virus type 1 gp41 by HIV-1 polypeptides expressed in Escherichia coli

Two monoclonal antibodies (MAbs) were produced in Balb/c mice by immunization with recombinant gp41 derived from expression of λ-BH10 cDNA of the human immunowdeficiency virus-1 (HIV-1) in the prokaryotic expression vector pEX-41 [1, 2]. Characterization of the epitopes recognized by these MAbs was done with HIV-1 envelope (env) fusion proteins expressed in Escherochia coli encoding ten distinct segments of the env proteins [3]. In comparison, another mouse MAb, M25 [4], a human MAb directed against gp41, which was produced by the xeno hydridoma line 3D6 [5, 6] and a pool of human patient sera containing antibodies to HIV-1 were tested. We were able to demonstrate that the epitopes recognized by our MAbs are located betweeni arg732 and ser759 [7] of the HIV-1 env glycoprotein gp160 of HTLV-III strain B. M25 reacted with epitopes between ser647 and pro731, which includes the hydrophobic transmembrane region of gp41 [4]. The human MAb against gp41, 3D6 [5, 6] reacts with epitopes between ile474 and trp646, a polypeptide stretch consisting of gp120 and gp41 specific amino acids. The human serum pool, positive for HIV-1 antibodies, reacted predominantly with antigenic determinants locatedp between ile474 and leu863. The recombinant env fusion proteins were initially produced to test the immunoreactivity with patient sera and to characterize epitopes which are relevant for immunodiagnostic purposes [3]. In this study, we showed that the set of recombinant evr proteins is also a simple and accurate tool for the characterization of MAbs directed to the HIV envelope proteins.

The noncompetitive blocker [3H]chlorpromazine labels three amino acids of the acetylcholine receptor gamma subunit: implications for the alpha-helical organization of regions MII and for the structure of the ion channel.

Labeling studies of Torpedo marmorata nicotinic acetylcholine receptor with the noncompetitive channel blocker [3H]chlorpromazine have led to the initial identification of amino acids plausibly participating to the walls of the ion channel on the alpha, beta, and delta subunits. We report here results obtained with the gamma subunit, which bring additional information on the structure of the channel. After photolabeling of the membrane-bound receptor under equilibrium conditions in the presence of agonist and with or without phencyclidine (a specific ligand for the high-affinity site for noncompetitive blockers), the purified labeled gamma subunit was digested with trypsin, and the resulting fragments were fractionated by HPLC. Sequence analysis of peptide mixtures containing various amounts of highly hydrophobic fragments showed that three amino acids are labeled by [3H]chlorpromazine in a phencyclidine-sensitive manner: Thr-253, Ser-257, and Leu-260. These residues all belong to the hydrophobic and putative transmembrane region MII of the gamma subunit. Their distribution along the sequence is consistent with an alpha-helical organization of this segment. The [3H]chlorpromazine-labeled amino acids are conserved at homologous positions in the known sequences of other ligand-gated ion channels and may, thus, play a critical role in ion-transport mechanisms.

The murine homologue of the T lymphocyte antigen CD28. Molecular cloning and cell surface expression.

The human T lymphocyte Ag CD28 (Tp44) is a homodimeric glycoprotein expressed on the surface of a majority of human peripheral T cells and thymocytes. Although exposure of T cells to anti-CD28 mAb does not activate T cells, stimulation of CD28 can synergize with signals transmitted through the TCR or other stimuli to augment proliferation and lymphokine production. We have used a portion of the human CD28 cDNA to isolate a homologous murine cDNA from an EL4 T lymphoma library. The murine clone has 61% nucleotide identity with the human cDNA. Both human and murine sequences exhibit homology with members of the Ig supergene family and CTLA-4, a T cell specific murine gene. Many characteristics of the human CD28 molecule are conserved within the putative murine CD28 polypeptide. The murine cDNA sequence encodes a polypeptide of 218 amino acids that has 68% identity with the human sequence. Both the murine and human molecules are integral membrane glycoproteins with hydrophobic signal peptide sequences and transmembrane region. All five potential N-linked glycosylation sites are conserved and six of the seven cysteine residues of the mouse protein are found in the human CD28 polypeptide. The murine cDNA is encoded by a single copy nonrearranging gene whose expression at the mRNA level is restricted to T cells. A rabbit antiserum was raised against a synthetic peptide corresponding to a hydrophilic portion of the translated murine cDNA sequence. This antiserum identifies an 80-kDa homodimer consisting of disulfide-bonded subunits of 40 kDa that is expressed on splenic T cells, thymocytes, and several T cell tumors, but not on B cells. deglycosylation studies indicate that four of the five N-linked glycosylation sites are used and that the mature core protein has a molecular mass of 25 kDa, close to that predicted by the cDNA sequence. Transfection of the murine cDNA into Chinese hamster ovary cells resulted in the expression of an 80-kDa dimeric molecule that was immunoprecipitated by the antipeptide antiserum. Taken together, these data provide strong support that we have identified the murine homologue of CD28.

Characterization and expression of the gene for the human Fc receptor gamma subunit. Definition of a new gene family.

The high affinity IgE receptor, Fc epsilon RI, is one of the key molecules involved in allergic reactions. It is a tetrameric complex (alpha beta gamma 2). The gamma chains from Fc epsilon RI are also subunits of other Fc receptors. We have isolated, characterized, and sequenced the gene for the human gamma chain of Fc epsilon RI. It consists of five exons and spans 4 kilobases. The leader sequence is encoded by two exons, the second of which also contains the short extracellular domain, the hydrophobic transmembrane region, and the beginning of the cytoplasmic tail. Three short exons encode the remaining of the polypeptide and the 3'-untranslated flanking sequence. The transcription initiation sites have been mapped. Comparison between the gene structures of the gamma chain of the Fc receptor and of the zeta chain of the T-cell receptor indicates that these genes have evolved from a common ancestor by duplication and that they define a new gene family. In addition to being localized on the same chromosome, both genes show an analogous organization of their exons. A high level of homology is found in three of their respective exons, and the splice sites between them are identical. Furthermore, gamma and zeta chains are essential for surface expression of their respective receptors. Therefore, gamma chains of Fc receptors and zeta chains of T-cell receptors may also define a new family of functionally related polypeptides. Expression studies in COS cells show that the human gamma chain alone is sufficient to achieve expression of the human alpha chain on the cell surface, whereas both beta and gamma chains are required for the surface expression of the rodent alpha chain.

Genomic organization of the human multidrug resistance (MDR1) gene and origin of P-glycoproteins.

The MDR1 gene, responsible for multidrug resistance in human cells, encodes a broad specificity efflux pump (P-glycoprotein). P-glycoprotein consists of two similar halves, each half including a hydrophobic transmembrane region and a nucleotide-binding domain. On the basis of sequence homology between the N-terminal and C-terminal halves of P-glycoprotein, we have previously suggested that this gene arose by duplication of a primordial gene. We have now determined the complete intron/exon structure of the MDR1 gene by direct sequencing of cosmid clones and enzymatic amplification of genomic DNA segments. The MDR1 gene includes 28 introns, 26 of which interrupt the protein-coding sequence. Although both halves of the protein-coding sequence are composed of approximately the same number of exons, only two intron pairs, both within the nucleotide-binding domains, are located at conserved positions in the two halves of the protein. The other introns occur at different locations in the two halves of the protein and in most cases interrupt the coding sequence at different positions relative to the open reading frame. These results suggest that the P-glycoprotein arose by fusion of genes for two related but independently evolved proteins rather than by internal duplication.

A point mutation in the neu oncogene mimics ligand induction of receptor aggregation.

The rat neu gene, which encodes a protein closely related to the epidermal growth factor receptor, is a proto-oncogene that can be converted into an oncogene by a point mutation. Both genes encode proteins with a relative molecular mass of 185,000 but the question of why the neu gene product, p185neu, is oncogenic, whereas the product of c-neu, p185c-neu, is not, remains unanswered. The proteins have several features common to the family of tyrosine kinase growth-factor receptors, including cysteine-rich external domains, a hydrophobic transmembrane region and a cytoplasmic tyrosine kinase domain. The oncogenic p185neu differs from p185c-neu by an amino-acid substitution in the transmembrane region of the glycoprotein: this replacement of valine by glutamic acid at position 664 induces increased intrinsic tyrosine kinase activity which is associated with transformation. Many glycoproteins with charged amino acids in the transmembrane region exist as multimeric complexes at the plasma membrane. We have therefore investigated the association state of both products of the neu gene and show that the oncoprotein p185neu is organized at the plasma membrane primarily in an aggregated form, but that p185c-neu is not. Induction of an aggregated state may mimic aspects of ligand-induced receptor aggregation resulting in enzymatic activation that leads to cellular transformation.

Molecular Cloning of CD1a (T6), a Human Epidermal Dendritic Cell Marker Related to Class I MHC Molecules

To investigate the structure, function, and control of CD1a, we have cloned a 1.6-kbp cDNA which encodes the expressed CD1a protein and includes untranslated 5' and 3' sequences and the poly-A tail. As the protein recognized by the monoclonal antibody OKT6, CD1a is a useful marker for Langerhans cells (LC). CD1a is found on these cells and on thymocytes, suggesting an important immunologic role for this molecule. We constructed a cDNA library in lambda gt10 using mRNA from MOLT-4, a cell line that expresses the CD1a surface antigen. We then screened the library with an oligonucleotide synthesized according to a known partial sequence for CD1a, and subcloned the cDNA and its restriction fragments into pGEM for sequencing and probe production. Based on this sequence the CD1a protein is predicted to consist of three extracellular domains (alpha 1-3), a hydrophobic transmembrane region, and a cytoplasmic tail. DNA 5' to the alpha 1 region may undergo alternative exon splicing. There is high sequence identity between the beta-2 microglobulin binding region of MHC I molecules and CD1a. The secondary structure predicted for CD1a is very similar to the actual structure of HLA-A2, a classical MHC I molecule. The similarity includes the beta pleated sheets and alpha helices which form the antigen binding groove of the alpha-1 and alpha-2 domains. The homology predicted between CD1a and HLA-A2 in these regions appears to exist on the level of secondary structure despite low primary nucleotide and amino acid sequence identity. The structural data and probes we have developed should facilitate studies of the function of CD1a as well as novel investigations of LC.

Molecular cloning and sequence determination of the fusion protein gene of human parainfluenza virus type 1

Undegraded mRNA transcripts were isolated from human parainfluenza virus type 1 (hPIV-1)-infected LLC-MK 2 cells and their size was determined through denaturing agarose electrophoresis. The two predominantly represented mRNA species (1.65 and 1.87 kb) are similar in size to other paramyxoviral mRNAs that encode their respective glycoproteins. The cDNA transcripts corresponding to these two mRNAs were used to construct two size-restricted cDNA libraries. A cDNA clone, containing a 1.87-kb insert, was identified as encoding the hPIV-1 fusion protein by positively hybridizing with a synthetic oligonucleotide mix whose sequence was derived from the conserved sequences of other paramyxoviral F 0 genes. The nucleotide sequence of the cDNA insert was determined and found to contain a single, large open reading frame encoding a putative protein of 60,795 Da consisting of 556 amino acids. Comparison of the amino acid sequence with the fusion proteins of other paramyxoviruses enabled the identification of the highly conserved amino acids of the F 1 N-terminus. In addition, the positions of the hydrophobic signal and transmembrane regions, cysteine, and proline residues are all conserved. These analyses confirm that the cDNA sequence is that of the F 0 protein. The 5′ end of the fusion protein mRNA was determined by primer extension to lie 155 bases beyond the 5′ end of the cDNA insert.

Primary structure of lymphocyte function-associated antigen 3 (LFA-3). The ligand of the T lymphocyte CD2 glycoprotein.

We have isolated the cDNA for human lymphocyte function-associated antigen 3 (LFA-3), the ligand of the T lymphocyte CD2 molecule. The identity of the clones was established by comparison of the deduced amino acid sequence to the LFA-3 NH2-terminal and tryptic peptide sequences. The cDNA defines a mature protein of 222 amino acids that structurally resembles typical membrane-anchored proteins. An extracellular domain with six N-linked glycosylation sites is followed by a hydrophobic putative transmembrane region and a short cytoplasmic domain. The mature glycoprotein is estimated to be 44-68% carbohydrate. Southern blots of human genomic DNA indicate that only one gene codes for human LFA-3. Northern blot analysis demonstrates that the LFA-3 mRNA of 1.3 kb is widely distributed in human tissues and cell lines.

Mutants of the Rous sarcoma virus envelope glycoprotein that lack the transmembrane anchor and cytoplasmic domains: analysis of intracellular transport and assembly into virions

The envelope glycoprotein complex of Rous sarcoma virus consists of a knoblike, receptor-binding gp85 polypeptide that is linked through disulfide bonds to a membrane-spanning gp37 spike. We used oligonucleotide-directed mutagenesis to assess the role of the hydrophobic transmembrane region and hydrophilic cytoplasmic domain of gp37 in intracellular transport and assembly into virions. Early termination codons were introduced on either side of the hydrophobic transmembrane region, and the mutated env genes were expressed from the late promoter of simian virus 40. This resulted in the synthesis of glycoprotein complexes composed of a normal gp85 and a truncated gp37 molecule that lacked the cytoplasmic domain alone or both the cytoplasmic and transmembrane domains. The biosynthesis and intracellular transport of the truncated proteins were not significantly different from those of the wild-type glycoproteins, suggesting that any protein signals for biosynthesis and intracellular transport of this viral glycoprotein complex must reside in its extracellular domain. The glycoprotein complex lacking the cytoplasmic domain of gp37 is stably expressed on the cell surface in a manner similar to that of the wild type. In contrast, the complex lacking both the transmembrane and cytoplasmic domains is secreted as a soluble molecule into the media. It can be concluded, therefore, that the transmembrane domain alone is essential for anchoring the RSV env complex in the cell membrane and that the cytoplasmic domain is not required for anchor function. Insertion of the mutated genes into an infectious proviral genome allowed us to assess the ability of the truncated gene products to be assembled into virions and to determine whether such virions were infectious. Viral genomes encoding the secreted glycoprotein were noninfectious, whereas those encoding a glycoprotein complex lacking only the cytoplasmic domain of gp37 were infectious. Virions produced from these mutant-infected cells contained normal levels of glycoprotein. The cytoplasmic tail of gp37 is thus not required for the assembly of envelope glycoproteins into virions. It is unlikely, therefore, that this region of gp37 interacts with viral core proteins during the selective incorporation of viral glycoproteins into the viral envelope.

Protein kinase activity of the insulin receptor.

The insulin receptor is an integral membrane glycoprotein (Mr approximately 300,000) composed of two alpha-subunits (Mr approximately 130,000) and two beta-subunits (Mr approximately 95,000) linked by disulphide bonds. This oligomeric structure divides the receptor into two functional domains such that alpha-subunits bind insulin and beta-subunits possess tyrosine kinase activity. The amino acid sequence deduced from cDNA of the single polypeptide chain precursor of human placental insulin receptor revealed that alpha- and beta-subunits consist of 735 and 620 residues, respectively. The alpha-subunit is hydrophilic, disulphide-bonded, glycosylated and probably extracellular. The beta-subunit consists of a short extracellular region which links the alpha-subunit through disulphide bridges, a hydrophobic transmembrane region and a longer cytoplasmic region which is structurally homologous with other tyrosine kinases like the src oncogene product and EGF receptor kinases. The cellular function of insulin receptors is dual: transmembrane signalling and endocytosis of hormone. The binding of insulin to its receptor on the cell membrane induces transfer of signal from extracellular to cytoplasmic receptor domains leading to activation of cell metabolism and growth. In addition, hormone-receptor complexes are internalized leading to intracellular proteolysis of insulin, whereas receptors are recycled to the membrane. These phenomena are kinetically well-characterized, but their molecular mechanisms remain obscure. Insulin receptor in different tissues and animal species are homologous in their structure and function, but show also significant differences regarding size of alpha-subunits, binding kinetics, insulin specificity and receptor-mediated degradation. We suggest that this heterogeneity of receptors may be linked to the diversity in insulin effects on metabolism and growth in various cell types. The purified insulin receptor phosphorylates its own beta-subunit and exogenous protein and peptide substrates on tyrosine residues, a reaction which is insulin-sensitive, Mn2+-dependent and specific for ATP. Tyrosine phosphorylation of the beta-subunit activates receptor kinase activity, and dephosphorylation with alkaline phosphatase deactivates the kinase. In intact cells or impure receptor preparations, a serine kinase is also activated by insulin. The cellular role of two kinase activities associated with the insulin receptor is not known, but we propose that the tyrosine- and serine-specific kinases mediate insulin actions on metabolism and growth either through dual-signalling or sequential pathways.(ABSTRACT TRUNCATED AT 400 WORDS)

The T cell differentiation antigen Leu-2/T8 is homologous to immunoglobulin and T cell receptor variable regions

Leu-2/T8 is a cell surface glycoprotein expressed by most cytotoxic and suppressor T lymphocytes. Its expression on T cells correlates best with recognition of class I major histocompatibility complex antigens, and it has been postulated to be a receptor for these proteins. We have determined the complete primary structure of Leu-2/T8 from the nucleotide sequence of its cDNA. The protein contains a classical signal peptide, two external domains, a hydrophobic transmembrane region, and a cytoplasmic tail. The N-terminal domain of the protein has striking homology to variable regions of immunoglobulins and the T cell receptor. The membrane-proximal domain appears to be a hinge-like region similar to that of immunoglobulin heavy chains. The superfamily of immunologically important surface molecules can now be extended to include Leu-2/T8.

The Drosophila EGF receptor gene homolog: Conservation of both hormone binding and kinase domains

Chicken v-erbB probe was used to isolate a unique clone of Drosophila melanogaster DNA. It maps by in situ hybridization to position 57F on chromosome 2. A complete nucleotide sequence of the coding region has been obtained. The putative Drosophila EGF receptor protein is similar in overall organization to the human homolog. It shows three distinct domains: an extracellular putative EGF binding domain, a hydrophobic transmembrane region, and a cytoplasmic kinase domain. The overall amino acid homology is 41% in the extracellular domain and 55% in the kinase domain. Two cysteine-rich regions, a hallmark of the human ligand-binding domain, have also been conserved. Fusion of the coding sequences of the kinase and extracellular domains generating the receptor gene must have occurred over 800 million years ago.

The complete sequence of the mRNA for the HLA-DR-associated invariant chain reveals a polypeptide with an unusual transmembrane polarity.

A non-polymorphic polypeptide is associated intracellularly with the alpha and beta chains of murine Ia antigens and of human HLA-DR antigens. The exact role and the structure of this invariant chain have not been determined so far. A cDNA clone encoding the 33 000 dalton human invariant chain has been isolated. The nucleotide sequence of a near full-length cDNA clone, together with the sequence of the 5' portion of the mRNA determined by primer-extension, are reported here. The protein structure deduced from that sequence shows an unusual feature: the presence of a hydrophobic transmembrane region near the NH2 terminus, and of two glycosylation sites near the middle, indicates that the invariant chain has a polarity of membrane insertion which is inverted relative to histocompatibility antigens and most transmembrane proteins.

Both alpha and beta chains of HLA-DC class II histocompatibility antigens display extensive polymorphism in their amino-terminal domains.

At least three class II antigens, all composed of an alpha and a beta subunit, are encoded in the human major histocompatibility complex, i.e., DR, DC and SB. Two cDNA clones, encoding a DC alpha and a DC beta chain, respectively, were isolated from a cDNA library of the lymphoblastoid cell line Raji (DR3,w6). The two polypeptides predicted from the nucleotide sequences of these clones are each composed of a signal peptide, two extracellular domains, a hydrophobic transmembrane region and a short cytoplasmic tail. Comparison of the DC alpha sequence with two previously published partial sequences shows that the majority of the differences is located in the amino-terminal domain. The differences are not randomly distributed; a cluster of replacements is present in the central portion of the amino-terminal domain. Likewise, the allelic polymorphism of the DC beta chains occurs preferentially in the amino-terminal domain, where three minor clusters of replacements can be discerned. The non-random distribution of the variability of DC alpha and beta chains may be due to phenotypic selection against replacement substitutions in the second domains of the polypeptides.

CDNA clone for the heavy chain of the human B cell alloantigen DC1: strong sequence homology to the HLA-DR heavy chain.

A cDNA library has been constructed from a B cell mRNA fraction enriched for HLA-DR sequences, and cDNA clones corresponding to sequences specifically expressed in B lymphocytes have been isolated by a differential screening procedure. Analysis of these clones with probes specific for the HLA-DR heavy chain gene allowed the characterization of HLA-DR heavy chain-related sequences. One clone, pDCH1, was demonstrated to encode the DC1 heavy chain because the amino acid sequence predicted from its nucleotide sequence matches eight out of nine residues available for comparison in the amino-terminal sequence of the DC1 heavy chain. The heavy chain of the DC1 alloantigen is composed of 232 amino acids and can be divided into two external domains, alpha 1 (amino acids 1-87) and alpha 2 (amino acids 88-181), a connecting peptide (amino acids 182-194), a hydrophobic transmembrane region (amino acids 195-217), and an intracytoplasmic region (amino acids 218-232). Comparison with the HLA-DR heavy chain reveals strong sequence homology in the second external Ig-like domain (alpha 2) and the transmembrane region. In contrast, the first external domain, the connecting peptide, and the intracytoplasmic region are less conserved.

CDNA clones for the heavy chain of HLA-DR antigens obtained after immunopurification of polysomes by monoclonal antibody.

A monoclonal antibody (HC 2.1) directed against the separated heavy chain of HLA-DR has been prepared. By binding HC 2.1 to polysomes from human B lymphoblastoid cells followed by the use of a protein A-Sepharose column as an immunoadsorbent, we have purified the mRNA coding for the HLA-DR heavy chain nearly to homogeneity. The immunopurified mRNA has been used to prepare labeled cDNA with which to probe cDNA libraries. Double-stranded cDNA was also made from the immunopurified mRNA and cloned directly into pBR322. Two clones, one from each of the above procedures, positively selected DR heavy chain message as assayed by cell-free translation and immunoprecipitation. One clone, pDRH-2 [500 base pairs plus 75 base pairs of poly(A)] contains the entire 3' untranslated region as well as coding information for the carboxy-terminal hydrophilic intracellular domain and part of the hydrophobic transmembrane region. Results of carboxypeptidase digestion of the heavy chains from detergent-solubilized (p34) and papain-treated (p33) HLA-DR antigen were consistent with the predicted protein sequence. Specific immunopurification of polysomes by defined monoclonal antibodies followed by direct cloning of cDNA to the highly purified mRNA is a powerful method for obtaining identified cDNA clones.

MRNA for surface immunoglobulin gamma chains encodes a highly conserved transmembrane sequence and a 28-residue intracellular domain.

To probe the structure of the gamma heavy chain of membrane IgG and the mRNA and gene segments that encode it, we have analyzed cDNA clones derived from a gamma 1 membrane RNA of B lymphoma 2PK-3. The nucleotide sequence of the clones indicated that membrane gamma 1 chains bear a COOH-terminal 71-residue segment that is absent from secretory gamma 1 chains. This terminus includes a 26-residue hydrophobic transmembrane region homologous to that of membrane mu chains and, significantly, a 28-residue intracellular domain found only on gamma chains. The extra domain suggests that receptor IgG, on memory B cells, may generate a different signal on binding antigen than does receptor IgM, on virgin B cells. The gamma 1 membrane terminus is encoded by two gene segments 1.5 and 2.4 kilobase pairs downstream from the C gamma 1 gene, and homologous segments occur 3' to the C gamma 2a and C gamma 3 genes. Small amounts of membrane gamma mRNAs persist in plasmacytomas secreting IgG1, IgG2a, or IgG2b, suggesting that competition between alternative RNA processing pathways governs the synthesis of membrane and secretory gamma chain mRNAs.