Mouse IgA allotypes have major differences in their hinge regions

Six IgA allotypes are serologically identifiable in inbred mice. The sequences of the PCR-amplified C alpha 1, C alpha 2 and C alpha 3 exons from the genomic DNA of mice of four previously unsequenced allotypes already have been compared with those of BALB/c and of a wild mouse, Mus pahari, in the literature. Sporadic differences, including several that may encode the known allotypic determinants, are found throughout the three exons, but major differences occur in the hinge. The hinge is longest in DBA/2 ( Igh-2(c)) mice, having an extra codon compared with that of BALB/c ( Igh-2(a)) and B10.A ( Igh-2(b)) mice. It is two codons shorter in CE ( Igh-2(f)) and four shorter in M. pahari, AKR and NZB (both Igh-2(d)) mice, but the position of the missing codons in the latter two strains is offset from that in M. pahari. The hinges in BALB/c ( Igh-2(a)) and DBA/2 ( Igh-2(c)) differ most from each other and from the other three allotypes, which are fairly closely related. Both BALB/c and DBA/2 have O-linked glycosylation sites, but they are in different positions in the hinge. Compared with BALB/c ( Igh-2(a)), B10.A( Igh-2(b)) has two extra Cys residues in the hinge, while DBA/2 ( Igh-2(c)), AKR/NZB ( Igh-2(d)) and CE ( Igh-2(f)) each have one. The differences in hinge length may have arisen by mismatching of highly repetitive portions of its sequence during meiotic recombination. Possible effects of the differences in hinge length and composition on the behavior of the mouse IgA allotypes are discussed.     

Immunoglobulin isoantigens (allotypes) in the mouse

Murine antisera raised against allogeneic lymphoid cells often contain antibodies to IgM allotypes. Rarely, allotypic antibodies to IgM have been found after immunization withB. pertussis anti-B. pertussis conjugates. Using both types of antibodies, we have defined a new constant-region locus for both secreted and membrane-bound chains. This locus,Ig-6, is closely linked to the previously described H-chain constant-region loci (Ig-1 throughIg-5) and is subject to allelic exclusion. We have identified three alleles and four antigenic specificities ofIg-6.Authors listed alphabetically     

Structural and genetic analysis of four chicken 7S immunoglobulin allotypes

Four 7S immunoglobulin allotypic specificities in three inbred chicken lines were demonstrated in two immunoglobulin regions, probably associated with the heavy chains. Two specificities were associated with papain-produced Fab fragments, most likely the Fd fragment since they were not demonstrated on the 17S immunoglobulin. The other allotypes were detected only on intact 7S immunoglobulin and were undetectable on the Fab and Fc fragments; therefore, they probably were associated with a region of the heavy chain sensitive to papain digestion. Among F₂ hybrids, these specificities segregated in a manner statistically indistinguishable from that expected of three codominant alleles at an autosomal locus. This locus was not closely associated with any of five chicken blood group loci.     

Structural and genomic correlates of hyperthermostability

While most organisms grow at temperatures ranging between 20 and 50 degrees C, many archaea and a few bacteria have been found capable of withstanding temperatures close to 100 degrees C, or beyond, such as Pyrococcus or Aquifex. Here we report the results of two independent large scale unbiased approaches to identify global protein properties correlating with an extreme thermophile lifestyle. First, we performed a comparative proteome analyses using 30 complete genome sequences from the three kingdoms. A large difference between the proportions of charged versus polar (noncharged) amino acids was found to be a signature of all hyperthermophilic organisms. Second, we analyzed the water accessible surfaces of 189 protein structures belonging to mesophiles or hyperthermophiles. We found that the surfaces of hyperthermophilic proteins exhibited the shift already observed at the genomic level, i.e. a proportion of solvent accessible charged residues strongly increased at the expense of polar residues. The biophysical requirements for the presence of charged residues at the protein surface, allowing protein stabilization through ion bonds, is therefore clearly imprinted and detectable in all genome sequences available to date.     

Mucosal immunity to influenza without IgA: an IgA knockout mouse model

IgA knockout mice (IgA-/-) were generated by gene targeting and were used to determine the role of IgA in protection against mucosal infection by influenza and the value of immunization for preferential induction of secretory IgA. Aerosol challenge of naive IgA-/- mice and their wild-type IgA+/+ littermates with sublethal and lethal doses of influenza virus resulted in similar levels of pulmonary virus infection and mortality. Intranasal and i.p. immunization with influenza vaccine plus cholera toxin/cholera toxin B induced significant mucosal and serum influenza hemagglutinin-specific IgA Abs in IgA+/+ (but not IgA-/-) mice as well as IgG and IgM Abs in both IgA-/- and IgA+/+ mice; both exhibited similar levels of pulmonary and nasal virus replication and mortality following a lethal influenza virus challenge. Monoclonal anti-hemagglutinin IgG1, IgG2a, IgM, and polymeric IgA Abs were equally effective in preventing influenza virus infection in IgA-/- mice. These results indicate that IgA is not required for prevention of influenza virus infection and disease. Indeed, while mucosal immunization for selective induction of IgA against influenza may constitute a useful approach for control of influenza and other respiratory viral infections, strategies that stimulate other Igs in addition may be more desirable.     

Structural correlates of imbibitional injury in Typha pollen

The ultrastructure of Typha latifolia pollen was examined as a function of pollen moisture content and incubation temperature, in order to identify possible lesions induced by imbibitional chilling. A syndrome of structural traits was found which characterizes damaged grains. Compared to viable grains, the protoplast of damaged pollen has a higher proportion of its volume occupied by vesicles, and less volume occupied by cytoplasm. Damaged grains also tend to have dilated cisternae of endoplasmic reticulum, larger starch grains and lipid bodies, poorly preserved mitochondria and membranes, and, sometimes, numerous electron-dense globules associated with membranes. The percentage of grains exhibiting this damage syndrome correlates closely with the number of ungerminated grains in most samples, regardless of moisture content or incubation temperature. Injury due to rapid imbibition from the dry state or to imbibitional chilling appear to be similar structurally, regardless of whether the stresses are imposed singly or together. The injury is not confined to one cell component (e.g., mitochondria), but may involve a generalized disruption of membranes. These results suggest that similar stress responses are elicited by imbibition from the dry state and by imbibitional chilling.     

Structural studies of human IgA1 and IgA2 immunoglobulins labeled with two different spin labels

The spin-label method was used for structural study of different subclasses of human immunoglobulin A. The spin label was incorporated into the protein part, as well as into carbohydrates of the IgA molecules. Well resolved outer wide extrema were characteristic of the ESR spectra of IgA spin-labeled at the protein moiety. ESR spectra of IgA tagged at carbohydrates reflected moderately immobilized rotation of the spin label. Dependencies of the parameters of ESR spectra of spin-labeled IgA1 and IgA2 upon viscosity at constant temperature have been investigated and a quantitative analysis of the isotherms was carried out. Spin-labeled oligosaccharide chains of IgA2 possessed great freedom of rotation. At least some of IgA1 oligosaccharides were closely attached to the protein moiety. Both proteins under study have shown flexible structure. The Fc fragment of IgA1 molecule appeared to have a rigid structure.     

Structural analysis of mouse S-antigen

Mouse S-antigen clones were isolated from a mouse retinal cDNA library using a bovine S-antigen cDNA probe. The largest clone (MSC-242) comprised 1532 bp and contained the entire coding sequence. The nucleotide sequence homology between the mouse and bovine coding regions was 84%, while non-coding regions appeared to be more divergent. The deduced amino acid sequence indicated that the mouse S-antigen had 403 residues and its molecular ratio was 44,930. An overall amino acid sequence similarity of 84% was observed between the mouse and bovine proteins. This degree of similarity dropped to 60% and 47% at the N and the C termini, respectively. The local homology with alpha-transducin observed in the bovine proteins, including the putative phosphoryl and rhodopsin binding sites, was conserved in the mouse as well. There was no overall sequence similarity with other proteins listed in the National Biomedical Research Foundation (NBRF) protein sequence database. Among the uveitopathogenic sites for experimental autoimmune uveitis (EAU), peptides N and M were identical to their bovine counterparts. Peptides 3 and K, however, were more divergent. The short repeats within these peptides were conserved.     

Structural and genetic studies on chicken 7S immunoglobulin allotypes. II. Distribution of allotypes on the 7S immunoglobulin of homozygous and heterozygous chickens.

We have previously reported that chicken 7S immunoglobulin (Ig) heavy (H) chain allotypes (CS-1 locus) segregate as phenogroups in F2 progeny. Specificity CS-1.1 formed a phenogroup with CS-1.4 in inbred chicken line UCD 2, and a second phenogroup with CS-1.3 in line UCD 3. To determine whether these phenogroups were formed by combinations of specificities on the same, or on separate subclasses of 7S Ig, their distribution on the 7S Ig molecules of birds homozygous for 7S Ig allotypes was analyzed by radioimmunoassay. Anti-CS-1.1 and anti-CS-1.3 alloantisera each bound more than 94% of line UCD 3 1252-7S Ig. Similar results were obtained with alloantisera to CS-1.1 and CS-1.4 WITH 125 I-7S Ig from line UCD 2. These results indicate that both phenogroups were formed by combinations of specificities present on the same H chain. Thus, each phenogroup described, probably is the product of a single structural gene which is responsible for more than 94% of the 7S Ig H chain constant regions. In F hybrids with the genotype CS-1.3, 1.3/CS-1.2, two populations of serum 7S Ig molecules were detected by direct and sequential binding analysis with specific alloantisera. One population of 7S Ig contained specificities CS-1.1 AND CS-1.3, but not CS-1.2; while the second population was exclusively the product of one parental allele. Consistent with a genetic regulatory mechanism involving allelic exclusion, no MS Ig containing allotypes produced by both alleles was detected.     

Rat kappa-chain allotypes

The presence of rat kappa-chain allotype specificities (RI-1a and b) has been studied in 13 subspecies of the seven native Australian species ofRattus. RI-la reactivity was not detected among these rats. On the other hand, extensive cross-reactivity was seen with RI-1b, some sera cross-reacting totally (R. leucopus cooktownensis), some not at all (R. colletti), and the remainder showing at least two distinct levels ofpartial cross-reactivity, confirming the existence of multiple specificities for Rl-lb. Three subspecies show polymorphism with respect to Rl-lb cross-reactivity (R. sordidus, R. colletti, andR. l. leucopus) and in one case (leucopus) breeding studies have indicated that there is allelic inheritance of this trait. The segregation of RI-1b reactivity has been studied in crosses and backcrosses made between species differing in their RI-1b reactivity, and the results are consistent with the existance of codominant alleles at a single locus. The fact that these species differ extensively in their karyotype opens the door to possible chromosomal localization of this and other genetic traits.     

Human immunoglobulin allotypes

More than twenty recombinant monoclonal antibodies are approved as therapeutics. Almost all of these are based on the whole IgG isotype format, but vary in the origin of the variable regions between mouse (chimeric), humanized mouse and fully human sequences; all of those with whole IgG format employ human constant region sequences. Currently, the opposing merits of the four IgG subclasses are considered with respect to the in vivo biological activities considered to be appropriate to the disease indication being treated. Human heavy chain genes also exhibit extensive structural polymorphism(s) and, being closely linked, are inherited as a haplotype. Polymorphisms (allotypes) within the IgG isotype were originally discovered and described using serological reagents derived from humans; demonstrating that allotypic variants can be immunogenic and provoke antibody responses as a result of allo-immunisation. The serologically defined allotypes differ widely within and between population groups; therefore, a mAb of a given allotype will, inevitably, be delivered to a cohort of patients homozygous for the alternative allotype. This publication reviews the serologically defined human IgG allotypes and considers the potential for allotype differences to contribute to or potentiate immunogenicity.     

Rat kappa-chain allotypes

The distribution of rat kappa-chain allotype specificities (RI-1a and 1b) was studied amongRattus rattus and a variety of other Asian rodents. No sera other than those ofRattus norvegicus showed the presence of RI-1a (DA type), whereas many cross-reacted with RI-1b (LEW-type). While manyR. rattus showedtotal cross-reactivity with RI-1b, various sera from the generaRattus, Bandicota, andTokudaia showed different levels ofpartial cross-reactivity. These results indicate that (1) anti-RI-1b reagents can detectmultiple specificities on LEW-type kappa chains, and (2) these RI-1b specificities, butnot RI-1a, are widely distributed among murid rodents, in seeming contradiction to amino acid sequence data suggesting that RI-1b is more closely related to ancestral rat kappa chains.