Distinct VH repertoires in primary and secondary B cell lymphocyte subsets in the preimmune repertoire of A/J mice: the CRI-A idiotype is preferentially associated with the HSA(low) B cell subset.

The anti-arsonate immune response of A/J mice is characterized by the occurrence of several recurrent idiotypes with a different temporal pattern of expression. The CRI-A idiotype is typically a memory idiotype since it appears late in the primary and dominates the secondary as well as subsequent immune responses. The CRI-C idiotype is present throughout the responses, including the primary one. Naive adult A/J mice treated repeatedly with anti-mu or anti-delta monoclonal antibodies exhibit a completely different balance of HSA(low) and HSA(high) B cell subsets and an opposite idiotype profile after immunization with p-azophenylarsonate coupled to hemocyanin. Anti-mu treatment leads to a striking enhancement of the HSA(low) cell subset associated with an earlier important synthesis of CRI-A(+) antibodies, while anti-delta treatment enhances significantly the HSA(high) compartment with a strong decrease of CRI-A and persistence of CRI-C1 antibodies. Semiquantitative PCR analysis reveals that the presence of CRI-A transcripts is associated with the HSA(low) compartment, while CRI-C transcripts are mainly associated with HSA(high) B cell subsets. This has been demonstrated with spleen cells of adult A/J mice treated with anti-mu or anti-delta antibodies and also with purified B cell subsets of unimmunized adult A/J mice and on neonatal spleen cells. It appears that the memory (CRI-A) idiotype is selected into the HSA(low) B cell subset before antigen arrival.

Molecular and cellular basis of the altered immune response against arsonate in irradiated A/J mice autologously reconstituted.

The humoral immune response to arsonate (Ars) in normal A/J mice is dominated in the late primary and particularly in the secondary response by a recurrent and dominant idiotype (CRIA) which is encoded by a single canonical combination of the variable gene segments: VHidcr11-DFL16.1-JH2 and Vkappa10-Jkappa1. Accumulation of somatic mutations within cells expressing this canonical combination or some less frequent Ig rearrangements results in the generation of high-affinity antibodies. By contrast, in partially shielded and irradiated A/J mice (autologous reconstitution) immunized with Ars-keyhole limpet hemocyanin (KLH), both the dominance of the CRIA idiotype and the affinity maturation are lost, whereas the anti-Ars antibody titer is not affected. To understand these alterations, we have analyzed a collection of 27 different anti-Ars hybridomas from nine partially shielded and irradiated A/J mice that had been immunized twice with Ars-KLH. Sequence analysis of the productively rearranged heavy chain variable region genes from those hybridomas revealed that (i) the canonical V(D)J combination was rare, (ii) the pattern of V(D)J gene usage rather corresponded to a primary repertoire with multiple gene combinations and (iii) the frequency of somatic mutations was low when compared to a normal secondary response to Ars. In addition, immunohistological analysis has shown a delay of 2 weeks in the appearance of full blown splenic germinal centers in autoreconstituting mice, as compared to controls. Such a model could be useful to understand the immunological defects found in patients transplanted with bone marrow.

An additional hypervariable region encoded by V gene segments occurs in Tcr V beta at a location compatible with its involvement in Tcr active site--a general model for alloreactivity.

The number of V alpha and V beta sequences of T cell receptors now available allows a meaningful analysis of their variability profiles. Variability plots were derived using a modified form of Wu and Kabat's algorithm: variability is not computed as a proportion of the number of different residues occurring at a position, but rather proportionally to the physicochemical differences between the different residues. Results show that the classical hypervariable regions occurring in immunoglobulins also occur in T cell receptors at equivalent positions. Contrary to immunoglobulins the framework of Tcr V regions displays many relatively variable regions and positions. This phenomenon can be connected with the genetic organization of V genes of T cell receptors which seem to avoid any framework homogenization and the resulting gene conversion. More importantly an additional hypervariable region was detected in V beta but not in V alpha. This fourth hypervariable region is located between the second and the D hypervariable CDR. The predicted three-dimensional location of this additional hypervariable region is compatible with a possible role in antigen recognition and therefore also in positive and/or negative selection. Furthermore our data suggest that this fourth hypervariable region is involved in the recognition of superantigens like bacterial enterotoxins. Indeed this additional hypervariable region is not detected when variability is derived using an alignment of the V beta subgroups stimulated by one toxin of S. aureus. Finally we propose a new and simple molecular model to explain alloreactivity as crossreactivity between the universe of shapes (isomers of conformation) of different MHC haplotypes.

Alpha secondary structures generate weak but recurrent periodicity in proteins.

Weak but recurrent periodic patterns characterize numerous actual proteins: the rate at which similarity occurs between residues i and i + 4 or i + 7 or i + 11 is very often slightly higher than predicted by chance. That result could indicate that numerous actual proteins derive from ancestral clearcut periodic sequences. Nevertheless, it is shown that this recurrent periodic pattern occurs much more significantly when analyses are restricted to the alpha-helical portions of proteins while it never occurs when beta-stranded subsequences are taken into account. This preferential location of periodicity inside protein subregions corresponding to alpha helices suggests that the recurrent pattern of weak periodicity could result from an ubiquitous physical property of alpha helices. The regular alternation of hydrophobicity, which is most often displayed by alpha helices, could then be the origin of weak periodicity.

Linear repetitions of amino acids and convergent evolution inside protein subregions of ordered secondary structures.

51 polypeptides of known 3-dimensional structures have been submitted to a search for internal similarities. It is shown that the frequency of proteins displaying significant amounts of internal similarities is higher than predicted by chance. A non-negligible part of those similarities probably occurs in connection with the existence of ordered secondary structures. Indeed, similarity occurs at a much more important rate when analyses are restricted to protein subsequences corresponding to alpha helices or beta pleated sheets. Furthermore, the correlation existing between the rates at which linear and inverted repeats occur inside protein subregions of ordered secondary structures suggests that a significant part of short similarities are analogies rather than homologies. An hypothesis is put forward suggesting that the regular alternations of hydrophobicity which characterize most of alpha helices and beta strands could provoke the occurrence of significant amounts of similarities inside protein sequences.

The sharing of amino acid short spans by ancestrally unrelated proteins may be the result of ubiquitous alpha and beta secondary structures.

Short homologies are often found when genetically unrelated proteins are compared but it is not known whether the rate at which they occur is or not above randomness. Comparing 190 pairs of unrelated proteins enable us to show that the frequency at which pairs of unrelated proteins share little spans of amino acids is compatible with chance. However, it appears that those short homologies are mainly located within protein subregions of identical secondary structure: the frequency at which pairs of unrelated proteins exhibit related spans of amino acids inside subregions of identical secondary structure is far above randomness. Those data suggest that the sharing of related spans of amino acids by genetically unrelated proteins could result from structural constraints imposed by the alpha or beta secondary structures.

Some thoughts on idiotypic networks and immunoregulation.

The first of two articles in which J. Urbain and C. Wuilmart discuss the manipulation of idiotype-anti-idiotype interactions in immune responses.

Relationships between alpha and beta secondary structures and amino-acid pseudosymmetrical arrangements.

A total of 51 polypeptides of known amino acid sequence and secondary structure have been screened for the presence of symmetrical arrangements of amino acids. Similarity between amino acids was derived by using a genetic test (minimum mutation distance) or a structural test (relative frequencies of amino acids substitutions in families of related proteins). It is shown that the frequency of proteins displaying symmetrical arrangements of amino acids is slightly higher than predicted by chance. In contrast, when the analysis is restricted to protein subregions displaying identical types of secondary structure, the frequency of proteins in which the alpha and beta subregions exhibit symmetrical arrangements of amino acids is significantly higher than predicted by chance. On the other hand, it is observed that more discriminatory results are always obtained when the structural test is used as a criterion for amino acid similarity. These data suggest that symmetrical arrangements of amino acids could result from structural constraints imposed either by the alpha or beta secondary structures. It is postulated that the regular alternation in hydrophobicity which is generally observed in the amino acid sub-sequences displaying alpha or beta secondary structures may be responsible for the occurrence of symmetrical arrangements of amino acids.

On the location of palindromes in immunoglobulin genes.

We present a statistical method for detection of palindromes in mRNA or DNA, starting from the protein sequence. Analysis of immunoglobulin genes by this method demonstrates that palindromic sequences are not randomly distributed. They are located at each side of the hypervariable regions in the variable (V) genes, whereas no such regular design is observed in the constant (C) genes. In addition, palindromic sequences overlap the V-C junction in all immunoglobulin classes and significant palindromes are present near residue 216 of the heavy chain, which is the end of deletions in many heavy chain diseases. The relevance of these palindromes to gene translocation and generation of diversity in antibodies is discussed.

Common origin and evolution of variable and constant regions of immunoglobulins.

Sequence data show that the immunoglobulins evolved from two sets of paralogous genes: a gene set coding for the V regions and another for the different C regions. A comparison of sequences from these two gene sets shows homology between the V and C sets of genes: this homology is only significant when VH is compared with Cmu1, Cmu2 and Cgamma1. There is a close agreement between our data drawn from sequence comparisons and the data of Poljak et al. (1974) drawn from crystallographic data. This finding is in agreement with the results of the phylogenetic trees of the C and V gene sets: they suggest that the VH subgroups and the first constant domain of the heavy chains are the most ancient. Moreover homology between the red blood cell glycophorin and Cmu2 suggests that immunoglobulins could have a common origin with some membrane proteins.

Linear and inverted repetitions in protein sequences.

An extensive search for internal regularities in amino acid sequences has been made, using both the genetic code and the relative frequencies of amino acid alternatives in homologous proteins. The two methods give very similar results and strongly suggest the occurrence of significant linear and inverted repetitions (similar sequences of opposite polarity) in several proteins. A hypothesis is developed to explain the occurrence of such internal regularities in proteins. This hypothesis is based on a process of duplication of an ancestral loop in which a symmetrical arrangement of amino acid allows stabilization by interaction between the amino acid side chains.