Search

Enduring forms of synaptic plasticity and memory require new protein synthesis, but little is known about the underlying regulatory mechanisms. Here, we investigate the role of MAPK signaling in these processes. Conditional expression of a dominant-negative form of MEK1 in the postnatal murine forebrain inhibited ERK activation and caused selective deficits in hippocampal memory retention and the translation-dependent, transcription-independent phase of hippocampal L-LTP. In hippocampal neurons, ERK inhibition blocked neuronal activity-induced translation as well as phosphorylation of the translation factors eIF4E, 4EBP1, and ribosomal protein S6. Correspondingly, protein synthesis and translation factor phosphorylation induced in control hippocampal slices by L-LTP-generating tetanization were significantly reduced in mutant slices. Translation factor phosphorylation induced in the control hippocampus by memory formation was similarly diminished in the mutant hippocampus. These results suggest a crucial role for translational control by MAPK signaling in long-lasting forms of synaptic plasticity and memory.

Calcineurin is a calcium-dependent protein phosphatase that has been implicated in various aspects of synaptic plasticity. By using conditional gene-targeting techniques, we created mice in which calcineurin activity is disrupted specifically in the adult forebrain. At hippocampal Schaffer collateral-CA1 synapses, LTD was significantly diminished, and there was a significant shift in the LTD/LTP modification threshold in mutant mice. Strikingly, although performance was normal in hippocampus-dependent reference memory tasks, including contextual fear conditioning and the Morris water maze, the mutant mice were impaired in hippocampus-dependent working and episodic-like memory tasks, including the delayed matching-to-place task and the radial maze task. Our results define a critical role for calcineurin in bidirectional synaptic plasticity and suggest a novel mechanistic distinction between working/episodic-like memory and reference memory.

Summary hypothesis is that an excess supply of target-derived neurotrophins should eliminate competition among thalMaturation of the visual cortex is influenced by visual amic inputs and therefore prevent the formation of ocuexperience during an early postnatal period. The fac- lar dominance columns or the physiological shift of ocutors that regulate such a critical period remain unclear. lar dominance following monocular deprivation (MD). We examined the maturation and plasticity of the vi- Indeed, intracortical administrations of BDNF or its neusual cortex in transgenic mice in which the postnatal tralizing agent, the trkB-IgG fusion protein, were shown rise of brain-derived neurotrophic factor (BDNF) was to alter the formation (Cabelli et al., 1995, 1997) and accelerated. In these mice, the maturation of GABAer- plasticity (Galuske et al., 1996) of ocular dominance colgic innervation and inhibition was accelerated. Fur- umns. Furthermore, cortical infusion of NT-4 rescued thermore, the age-dependent decline of cortical long- LGN neurons from the effect of MD (Riddle et al., 1995). term potentiation induced by white matter stimulation, These results are consistent with the notion that a ligand a form of synaptic plasticity sensitive to cortical inhibi- of TrkB, either BDNF and/or NT-4, acts directly on LGN tion, occurred earlier. Finally, transgenic mice showed axons and is involved in their activity-dependent compea precocious development of visual acuity and an ear- tition for layer IV synaptic targets during ocular domilier termination of the critical period for ocular domi- nance segregation. However, alternative interpretations nance plasticity. We propose that BDNF promotes the for these findings are plausible. For example, neuromaturation of cortical inhibition during early postnatal trophins could affect LGN axons indirectly, via alterlife, thereby regulating the critical period for visual ations in the dendritic morphology of cortical neurons, cortical plasticity.

To investigate the role of synaptic plasticity in the place-specific firing of the hippocampus, we have applied multiple electrode recording techniques to freely behaving mice with a CA1 pyramidal cell–specific knockout of the NMDAR1 gene. We have discovered that although the CA1 pyramidal cells of these mice retain place-related activity, there is a significant decrease in the spatial specificity of individual place fields. We have also found a striking deficit in the coordinated firing of pairs of neurons tuned to similar spatial locations. Pairs have uncorrelated firing even if their fields overlap. These results demonstrate that NMDA receptor–mediated synaptic plasticity is necessary for the proper representation of space in the CA1 region of the hippocampus.

The brain dopaminergic system is a critical modulator of basal ganglia function and plasticity. To investigate the contribution of the dopamine D1 receptor to this modulation, we have used gene targeting technology to generate D1 receptor mutant mice. Histological analyses suggested that there are no major changes in general anatomy of the mutant mouse brains, but indicated that the expression of dynorphin is greatly reduced in the striatum and related regions of the basal ganglia. The mutant mice do not respond to the stimulant and suppressive effects of D1 receptor agonists and antagonists, respectively, and they exhibit locomotor hyperactivity. These results suggest that the D1 receptor regulates the neurochemical architecture of the striatum and is critical for the normal expression of motor activity.

mGluR1 mutant mice are viable but show characteristic cerebellar symptoms such as ataxic gait and intention tremor. The anatomy of the cerebellum is not overtly disturbed. Excitatory synaptic transmission from parallel fibers (PFs) to Purkinje cells and that from climbing fibers (CFs) to Purkinje cells appear to be functional, and voltage-gated Ca2+ channels of Purkinje cells are normal. Both PF and CF synapses display normal short-term synaptic plasticity to paired stimuli. By marked contrast, long-term depression (LTD) is clearly deficient and conditioned eyeblink response is impaired. We conclude that mGluR1 is required for the induction of LTD and that the ataxic behavior and impaired eyeblink conditioning of the mGluR1 mutant mice are primarily due to deficient LTD.

Positive and negative selection of a lymphocytic choriomeningitis virus (LCMV) peptide-specific, H-2Db-restricted T cell clone (P14) was studied using TAP1- and TAP1+ mice transgenic for P14 T cell receptor (TCR) alpha and beta genes. Positive selection of transgenic CD8+ P14 cells was impaired in TAP1- mice. Addition of the LCMV peptide to TAP1- fetal thymic organ cultures (FTOCs) at low and high concentrations induced positive and negative selection of CD8+ P14 cells, respectively, while addition of the same peptide to TAP1+ FTOCs induced negative selection even at low concentrations. Both types of selection were peptide specific. Thus, a critical parameter that controls the fate of a thymocyte seems to be the number of TCRs engaged with complexes of peptide and major histocompatibility complex. When this number is low, positive selection occurs, and when it is high, negative selection takes place. These findings support a differential avidity model of T cell selection.

Calcium-phospholipid-dependent protein kinase (PKC) has long been suggested to play an important role in modulating synaptic efficacy. We have created a strain of mice that lacks the γ subtype of PKC to evaluate the significance of this brain-specific PKC isozyme in synaptic plasticity. Mutant mice are viable, develop normally, and have synaptic transmission that is indistinguishable from wild-type mice. Long-term potentiation (LTP), however, is greatly diminished in mutant animals, while two other forms of synaptic plasticity, long-term depression and paired-pulse facilitation, are normal. Surprisingly, when tetanus to evoke LTP was preceded by a low frequency stimulation, mutant animals displayed apparently normal LTP. We propose that PKCγ is not part of the molecular machinery that produces LTP but is a key regulatory component.

We are undertaking a genetic approach to investigate the role that synaptic modulation in the mammalian central nervous system plays in learning and memory and to identify relevant molecular components. We have generated mice deficient in the gamma isoform of protein kinase C (PKC gamma), an enzyme that has previously been implicated in both long-term potentiation (LTP) and learning and memory. These mice have a modified LTP of synaptic transmission in the hippocampus. We demonstrate that PKC gamma-mutant mice can learn to carry out hippocampus-dependent tasks, although mild deficits are evident. Thus, hippocampal CA1 LTP induced by the conventional tetanic stimulation is not essential for the mice to exhibit spatial and contextual learning. Furthermore, the modification of hippocampal synaptic plasticity correlates with the learning deficits we observe.

We describe the spontaneous development of inflammatory bowel disease (IBD) in several immunodeficient mouse strains created via gene targeting in embryonic stem cells. Chronic colitis was observed in T cell receptor (TCR) alpha mutant, TCR beta mutant, TCR beta x delta double mutant, or class II major histocompatibility complex (MHC) mutant mice, but not in recombination-activating gene RAG-1 mutant mice or nude mice kept in the same specific pathogen-free animal facility. This clinical pattern suggests that the disease requires the presence of B lymphocytes and the absence of class II MHC-restricted CD4+ alpha beta T cells. IBD in the mutant mice has some of the features of the human disease ulcerative colitis. Based on these results, we suggest that dysfunction of the mucosal immune system may underly the pathogenesis of some types of IBD in humans.

T cells detect infection of cells by recognizing peptide fragments of foreign proteins bound to class I molecules of the major histocompatibility complex (MHC) on the surface of the infected cell. MHC class I molecules bind peptide in the endoplasmic reticulum, and analysis of mutant cells has demonstrated that an adequate supply of peptides requires the presence of two genes in the MHC class II locus that encode proteins called transporters associated with antigen processing (TAP) 1 and 2. TAP1 and TAP2 are members of the ATP-binding cassette family of membrane translocators. In this study, we demonstrate in a cell-free system that TAP1 is part of an ATP-dependent, sequence-specific, peptide translocator.

Mice deficient in the gene encoding the peptide transporter associated with antigen processing (TAP1) have drastically reduced levels of surface expression of MHC class I molecules and few CD8+ T cells. Addition of class I binding peptides to cultured fetal thymi from TAP1 mutant mice invariably allowed the rescue of class I expression, while only some of these peptides promoted the positive selection of CD8+ T cells, which were polyclonal and specific for the peptide-MHC complex. A nonselecting peptide was converted to a mixture of selecting peptides when the residues involved in the interaction with TCRs were altered. A mixture of self-peptides derived from C57BL/6 thymi induced positive selection of CD8+ T cells at concentrations that gave relatively low class I surface expression. The implication of these observations is that self-peptides determine, in part, the repertoire of specificities exhibited by CD8+ T cells.

T cells bearing T cell receptor (TCR) gamma and delta chain heterodimers are first generated early in ontogeny. They form distinct subsets that differ in their TCR repertoires and tissue distribution. Disruption of the mouse TCR C delta gene segment by a gene targeting method caused the complete loss of T cells bearing TCR gamma delta chains, but had little or no effect on the development of T cells bearing TCR alpha beta chains. The analyses of TCR gamma and delta genes in the mutant mice suggest that intracellular mechanisms acting at the level of DNA rearrangement play key roles in the differential gamma and delta gene rearrangements and in the generation of the highly restricted junctional sequences during fetal thymic development.

Optogenetic technologies employ light to control biological processes within targeted cells in vivo with high temporal precision. Here, we show that application of molecular trafficking principles can expand the optogenetic repertoire along several long-sought dimensions. Subcellular and transcellular trafficking strategies now permit (1) optical regulation at the far-red/infrared border and extension of optogenetic control across the entire visible spectrum, (2) increased potency of optical inhibition without increased light power requirement (nanoampere-scale chloride-mediated photocurrents that maintain the light sensitivity and reversible, step-like kinetic stability of earlier tools), and (3) generalizable strategies for targeting cells based not only on genetic identity, but also on morphology and tissue topology, to allow versatile targeting when promoters are not known or in genetically intractable organisms. Together, these results illustrate use of cell-biological principles to enable expansion of the versatile fast optogenetic technologies suitable for intact-systems biology and behavior.

The transporter associated with the antigen processing 1 (TAP1) gene encodes a subunit for a transporter, presumed to be involved in the delivery of peptides across the endoplasmic reticulum membrane to class I molecules. We have generated mice with a disrupted TAP1 gene using embryonic stem cell technology. TAP1-deficient mice are defective in the stable assembly and intracellular transport of class I molecules and consequently show severely reduced levels of surface class I molecules. These properties are strikingly similar to those described for the TAP2 mutant cell line RMA-S. Cells from the TAP1-deficient mice are unable to present cytosolic antigens to class I-restricted cytotoxic T cells. As predicted from the near absence of class I surface expression, TAP1-deficient mice lack CD4-8+ T cells.

The gamma delta T cell receptor (TCR) derived from the mouse KN6 T cell hybridoma recognizes an autologous determinant encoded by a broadly expressed gene mapping in the TL region of the major histocompatibility complex (MHC). We have cloned the gene and demonstrated that it is a novel class I gene (designated 27b) belonging to a hitherto undescribed TL region gene cluster in strain C57BL/6. The BALB/c allele of 27b, gene T17c, is defective because it lacks an appropriate splice acceptor site, which explains the lack of recognition of BALB/c stimulator cells by the KN6 cells. We propose that gamma delta TCR and nonclassical MHC and MHC-related class I molecules have coevolved to recognize a conserved set of endogenous and foreign determinants.

SummaryCortical representations of visual information are modified by an animal's visual experience. To investigate the mechanisms in mice, we replaced the coding part of the neural activity-regulated immediate early gene Arc with a GFP gene and repeatedly monitored visual experience-induced GFP expression in adult primary visual cortex by in vivo two-photon microscopy. In Arc-positive GFP heterozygous mice, the pattern of GFP-positive cells exhibited orientation specificity. Daily presentations of the same stimulus led to the reactivation of a progressively smaller population with greater reactivation reliability. This adaptation process was not affected by the lack of Arc in GFP homozygous mice. However, the number of GFP-positive cells with low orientation specificity was greater, and the average spike tuning curve was broader in the adult homozygous compared to heterozygous or wild-type mice. These results suggest a physiological function of Arc in enhancing the overall orientation specificity of visual cortical neurons during the post-eye-opening life of an animal.

Using the phage P1–derived Cre/loxP recombination system, we have developed a method to create mice in which the deletion (knockout) of virtually any gene of interest is restricted to a subregion or a specific cell type in the brain such as the pyramidal cells of the hippocampal CA1 region. The Cre/loxP recombination–based gene deletion appears to require a certain level of Cre protein expression. The brain subregional restricted gene knockout should allow a more precise analysis of the impact of a gene mutation on animal behaviors.

The brain mesoaccumbens dopamine system is intricately involved in the psychomotor stimulant activities of cocaine. However, the extent to which different dopamine receptors mediate these effects has not yet been firmly established. The present study used dopamine D1 receptor mutant mice produced by gene targeting to investigate the role of this receptor in the effects induced by cocaine. In contrast with wild-type mice, which showed a dose-dependent increase in locomotion, D1 mutant mice exhibited a dose-dependent decrease. Electrophysiological studies of dopamine-sensitive nucleus accumbens neurons demonstrated a marked reduction in the inhibitory effects of cocaine on the generation of action potentials. In addition, the inhibitory effects of dopamine as well as D1 and D2 agonists were almost completely abolished, whereas those of serotonin were unaffected. D2-like dopamine receptor binding was also normal. These results demonstrate the essential role of the D1 receptor in the locomotor stimulant effects of cocaine and in dopamine-mediated neurophysiological effects within the nucleus accumbens

We generated a novel strain of mutant mouse with a deletion in the gene encoding metabotropic glutamate receptor 1 (mGluR1). Gross anatomy of the hippocampus, excitatory synaptic transmission, long-term depression, and short-term potentiation in the hippocampal CA1 region are all apparently normal in the mutant mice. In contrast, long-term potentiation (LTP) is substantially reduced, and a moderate level of impairment is observed in context-specific associative learning. We propose that mGluR1 is not "in line" in LTP production, but rather modulates the plasticity process, and hence affects context-specific associative learning.

We have generated TCR transgenic mice (T/R+) specific for myelin basic protein (MBP) and crossed them to RAG-1-deficient mice to obtain mice (T/R-) that have T cells expressing the transgenic TCR but no other lymphocytes. Both T/R+ and T/R- mice carry, in the lymph nodes and spleen, large numbers of the potentially encephalitogenic CD4+ anti-MBP T cells. These cells respond to MBP in vitro but show no signs of activation in vivo. Nevertheless, approximately 14% of H-2u T/R+ and 100% of H-2u T/R- mice developed spontaneous experimental autoimmune encephalomyelitis (EAE) within 12 months. These data indicate that EAE can be mediated by CD4+ anti-MBP T cells in the absence of any other lymphocytes and that nontransgenic lymphocytes that are present in T/R+ but absent in T/R- mice have a protective effect. The data also suggest that spontaneous EAE may be triggered by an in situ activation of CD4+ anti-MBP cells in the nervous system.

Sensory pathways of the brain generally develop from crudely wired networks to precisely organized systems. Several studies have implicated neural activity-dependent mechanisms, including N-methyl-D-aspartate (NMDA) receptors, in this refinement process. We applied the gene targeting to the NMDAR1 gene and created a mutant mouse that lacks functional NMDA receptors. The development of whisker-related patterns in the trigeminal nuclei of the mutant mice and their normal littermates was compared. We show that in the mutant mice pathfinding, initial targeting, and crude topographic projection of trigeminal axons in the brainstem are unaffected, but that whisker-specific patches fail to form. Our results provide a direct demonstration of the involvement of the NMDA receptor in the formation of periphery-related neural patterns in the mammalian brain.

The V(D)J recombination activation gene RAG-1 was isolated on the basis of its ability to activate V(D)J recombination on an artificial substrate in fibroblasts. This property and the expression pattern in tissues and cell lines indicate that RAG-1 either activates or catalyzes the V(D)J recombination reaction of immunoglobulin and T cell receptor genes. We here describe the introduction of a mutation in RAG-1 into the germline of mice via gene targeting in embryonic stem cells. RAG-1-deficient mice have small lymphoid organs that do not contain mature B and T lymphocytes. The arrest of B and T cell differentiation occurs at an early stage and correlates with the inability to perform V(D)J recombination. The immune system of the RAG-1 mutant mice can be described as that of nonleaky scid mice. Although RAG-1 expression has been reported in the central nervous system of the mouse, no obvious neuroanatomical or behavioral abnormalities have been found in the RAG-1-deficient mice.

Using a pCRI plasmid containing an enzymatically synthesized, full-length DNA transcript of immunoglobulin lambda chain mRNA as the hybridization probe in the Southern gel blotting experiments (Southern, 1975), we identified three DNA fragments of 8.6, 4.8 and 3.5 kb in Eco RI-digested total DNA from BALB/c mouse embryos. A fourth fragment of 7.4 kb was found in addition to these three fragments in similarly digested total DNA from a lambda chain-secreting myeloma (HOPC 2020). We have cloned the four DNA fragments in an EK-2 phage vector, lambdaWES, and characterized them with respect to size, type of lambda gene sequences contained and position of these sequences in the fragments, using agarose gel electrophoresis, the gel blotting technique and electron microscopic R loop mapping. The embryonic DNA clones Ig 99 lambda, Ig 25lambda and Ig 13lambda contain one copy each of V lambdaI, C lambdaI and V lambdaII sequences, respectively, while the myeloma DNA clone Ig 303lambda contains one copy each of V lambdaI and C lambdaI sequences that are separated by a 1.2 kb nontranslated DNA segment. Ig 25lambda was also shown to contain a DNA segment of approximately 40 base pairs (bp) (J sequence) that lies 1.2 kb away from the C lambdaI sequence and is homologous to the V-C junction region of a lambdaI mRNA. Heteroduplex analysis of the three lambdaI DNA clones revealed that Ig 303lambda DNA is composed of two parts, one of which is entirely homologous to one end of Ig 99lambda, and the other to one end of Ig 25lambda DNA. The sequence arrangement observed in the cloned DNA is the same as that in the corresponding cellular DNA. This was shown by identifying certain restriction enzyme cleavage sites on the cloned DNAs and demonstrating the presence of these sites in the total cellular DNA by the gel blotting technique. The site of the homology switch is at the boundary of the V sequence and the 1.2 kb nontranslated DNA segment, and corresponds to the position of the J sequence on the Ig 25lambda DNA. We consider the above experimental results the most direct evidence for somatic rearrangement in immunoglobulin genes. We discuss the significance of these findings for the origin of genes in the evolution of higher organisms and in cell differentiation.

Nucleotide sequences of a large number of V-(D)-J junctions of T cell receptor (TCR) γ and δ genes show that most fetal thymocytes express on their surface one of just two γδ TCRs known to be expressed by epidermal γδ T cells (s-IEL) or intraepithelial γδ T cells associated with female reproductive organs (r-IEL). In contrast, γδ TCRs expressed on adult thymocytes are highly diverse as a result of multiple combinations of gene segments as well as junctional deletions and insertions, indicating that developmental time- and cell lineage-dependent mechanisms exist that control the extent of γδ TCR diversity. In addition, this study revealed a new type of junctional insertion (P nucleotides), which led to a new model of V-(D)-J joining generally applicable to immunoglobulin and TCR genes.

We present the initial characterization of a novel family of genes that rearrange in T cells, but do not encode either of the defined (alpha/beta) subunits of the clone-specific heterodimer of the T cell receptor. The family comprises at least three variable (V) gene segments, three constant (C) gene segments, and three junction (J) gene segments. In a cloned cytolytic T lymphocyte, 2C, one of each of these fragments has productively rearranged to yield an expressed VJC transcription unit, which shows no evidence for somatic mutation. Short sequences similar to those implicated in immunoglobulin gene and T cell receptor beta chain gene rearrangement flank the V and J segments of this family. The linkage of two of the three V gene segments has been determined: the segments lie approximately 2.5 kb apart, and are arranged head-to-head. The inverted arrangement may cast light upon the mechanisms utilized by lymphocytes for gene rearrangement.

Summary We have produced a mouse strain in which the deletion of the NMDAR1 gene is restricted to the CA1 pyramidal cells of the hippocampus by using a new and general method that allows CA1-restricted gene knockout. The mutant mice grow into adulthood without obvious abnormalities. Adult mice lack NMDA receptor–mediated synaptic currents and long-term potentiation in the CA1 synapses and exhibit impaired spatial memory but unimpaired nonspatial learning. Our results strongly suggest that activity-dependent modifications of CA1 synapses, mediated by NMDA receptors, play an essential role in the acquisition of spatial memories.

It is generally believed that a smooth execution of a compound movement, or motor coordination, requires learning of component movements as well as experience-based refinement of the motor program as a whole. PKCγ mutant mice display impaired motor coordination but intact eyeblink conditioning, a form of component movement learning. Cerebellar long-term depression, a putative cellular mechanism for component motor learning, is also unimpaired. Thus, PKCγ mutant mice are defective in refinement of the motor program. In the accompanying paper, we demonstrate that innervation of multiple climbing fibers onto Purkinje cells persists in adulthood in these mutant mice. We propose that this defective elimination of surplus climbing fibers underlies motor discoordination.

Presenilin-1(PS1) is the major gene responsible for early-onset familial Alzheimer's disease (FAD). To understand the normal function of PS1, we have generated a targeted null mutation in the murine homolog of PS1. We report that PS1−/− mice die shortly after natural birth or Caesarean section. The skeleton of homozygous mutants is grossly deformed. Hemorrhages occur in the CNS of PS1 null mutants with varying location, severity, and time of onset. The ventricular zone of PS1−/− brains is markedly thinner by embryonic day 14.5, indicating an impairment in neurogenesis. Bilateral cerebral cavitation caused by massive neuronal loss in specific subregions of the mutant brain is prominent after embryonic day 16.5. These results show that PS1 is required for proper formation of the axial skeleton, normal neurogenesis, and neuronal survival.

Neuronal oscillations have been hypothesized to play an important role in cognition and its ensuing behavior, but evidence that links a specific neuronal oscillation to a discrete cognitive event is largely lacking. We measured neuronal activity in the entorhinal-hippocampal circuit while mice performed a reward-based spatial working memory task. During the memory retention period, a transient burst of high gamma synchronization preceded an animal’s correct choice in both prospective planning and retrospective mistake correction, but not an animal’s incorrect choice. Optogenetic inhibition of the circuit targeted to the choice point area resulted in a coordinated reduction in both high gamma synchrony and correct execution of a working-memory-guided behavior. These findings suggest that transient high gamma synchrony contributes to the successful execution of spatial working memory. Furthermore, our data are consistent with an association between transient high gamma synchrony and explicit awareness of the working memory content., RIKEN Brain Science Institute, Picower Institute for Learning and Memory (Innovation Fund), Human Frontier Science Program (Strasbourg, France) (Fellowship)

We have studied the DNA sequences required for high level expression of a cloned heavy chain immunoglobulin gene stably introduced into mouse myeloma cells by DNA transfection. We found that DNA sequences derived from the germ line JH-C mu region are required for accurate and efficient transcription from a functionally rearranged VH promoter. Similar to viral transcriptional enhancer elements, these cellular sequences stimulate transcription from either the homologous VH gene segment promoter or a heterologous SV40 promoter. They are active when placed on the 5' or 3' side of the rearranged VH gene segment and they function when their orientation is reversed. However, unlike viral enhancers, the Ig gene enhancer appears to act in a tissue-specific manner, since it is active in mouse B cells but not in mouse fibroblasts. The nucleotide sequence of the Ig enhancer region contains repeating elements that closely resemble sequence the possible role of tissue-specific transcription in cell differentiation and malignant transformation.

We have determined the nucleotide sequences of the germ line gene as well as a corresponding somatically mutated and rearranged gene coding for a mouse immunoglobulin lambdaI type light chain. These sequencing studies were carried out on three Eco RI-DNA fragments which had been cloned from BALB/c mouse embryos or a lambdaI chainsecreting myeloma, H2020. The embryonic DNA clone Ig 99lambda contains two protein-encoding segments, one for the majority of the hydrophobic leader (L) and the other for the rest of the leader and the variable (V) region of the lambda0 chain (Cohn et al., 1974); these segments are separated by a 93 base pair (bp) intervening sequence (I-small). The coding of the V region ends with His at residue 97. The second embryonic DNA clone Ig 25lambda includes a 39 bp DNA segment (J) coding for the rest of the conventionally defined V region (that is, up to residue 110), and also contains the sequence coding for the constant (C) region approximately 1250 untranslated bp (I-large) away from the J sequence. The J sequence is directly linked with the V-coding sequence in the myeloma DNA clone, Ig 303lambda, which has the various DNA segments arranged in the following order: 5' untranslated region, L, l-small, V linked with J, l-large, C, 3' untranslated sequence. The lg 303lambda V DNA sequence codes for the V region synthesized by the H2020 myeloma and is different from the lg 99lambda V DNA sequence by only two bases. No silent base change was observed between the two DNA clones for the entire sequence spanning the 5' untranslated regions and the V-coding segments. These results confirm the previously drawn conclusion that an active complete lambdaI gene arises by somatic recombination that takes place at the ends of the V-coding DNA segment and the J sequence. No sequence homology was observed at or near the sites of the recombination.