Zoltán Lóránt Nagy

A hemagglutinin-based multipeptide construct elicits enhanced protective immune response in mice against influenza A virus infection

Multipeptide constructs, comprising adjacent sequences of the 317-341 intersubunit region of immature influenza A hemagglutinin (H1N1), were designed and the functional properties of these branched peptides were compared to that of the corresponding linear peptides. In vivo studies revealed that the immunogenicity of the peptides was dependent on the presence of the hydrophobic fusion peptide (comprised in FP3), encompassing the N-terminal 1-13 sequence of the HA2 subunit. Antibody and T cell recognition, however, was directed against the 317-329 HA1 sequence, comprised in the P4 peptide. Multiple copies of P4, covalently linked by branched lysine residues, significantly enhanced the efficiency of antibody binding and the capacity of peptides to elicit B- and T-cell responses. A fraction of peptide induced antibodies reacted with immature or with proteolitically cleaved hemagglutinin (HA) molecules pretreated at low pH. Immunization with a multipeptide construct, (P4)4-FP3, not only resulted in elevated antibody and T cell responses but conferred enhanced protection against lethal A/PR/8/34 (H1N1) infection as compared to its subunit peptides. The beneficial functional properties of this artificial peptide antigen may be acquired by multiple properties including: (i) stabilized peptide conformation which promotes strong, polyvalent binding to both antibodies and MHC class II molecules; (ii) appropriate P4 conformation for antibody recognition stabilized by the covalently coupled fusion peptide, resulting in the production of virus cross reactive antibodies which inhibit the fusion activity of the virus; (iii) activation of peptide specific B cells which potentiate antigen presentation and peptide specific T cell responses; and (iv) generation of helper T cells which secrete lymphokines active in the resolution of infection.

T cell recognition of the posttranslationally cleaved intersubunit region of influenza virus hemagglutinin

The influenza virus hemagglutinin is synthesized as a single polypeptide chain, but upon maturation it will posttranslationally be modified by a host cell related trypsin-like enzyme. The enzymatic cleavage attacks the so-called intersubunit region of the molecule giving rise to covalently linked HA1 and HA2 subunits. An I-Ed-restricted T cell epitope was identified in the highly conserved intact intersubunit region of the influenza virus hemagglutinin. T cell recognition of a 25-mer synthetic peptide comprising the intact intersubunit region does not require further processing and the elimination of the intervening Arg residue coupling the fusion peptide to the C-terminal segment of HA1 does not abolish the T cell activating capacity. The fine specificity pattern of a T cell hybridoma similar to that of the polyclonal T cell response demonstrates that a single T cell receptor is able to recognize peptides of different sizes representing not only the uncleaved but also the cleaved form of this hemagglutinin region. Based on specificity studies the epitope was localized to the C-terminal 11 amino acids of the HA1 subunit. The cross-reactivity of peptide-primed T cells with influenza virus infected antigen-presenting cells shows that fragments comprising the identified epitope of the intersubunit region can be generated as a result of natural processing of the hemagglutinin molecule. As antigen-presenting cells are lacking the enzyme which is responsible for the posttranslational modification of newly synthesized hemagglutinin molecules, the role of immature viral proteins in immune recognition is discussed.

The Intersubunit Region of the Influenza Virus Haemagglutinin is Recognized by Antibodies During Infection

The influenza virus haemagglutinin has an important role in the infectious cycle of the virus and carries multiple B and T cell epitopes. It is synthesized as a single polypeptide chain but viral infectivity depends on its post‐translational enzymatic cleavage. The cleavage site of a trypsin‐like enzyme responsible for this modification is found in the most conserved intersubunit region of the molecule. In this study the role of this region in antibody recognition was investigated. Synthetic peptides comprising the intact and cleaved forms of the intersubunit segment were used to examine the specificity of virus‐ or peptide‐ induced antibodies. The immune response elicited by viral infection resulted in the appearance of antibodies capable of neutralizing the virus without interfering with its binding to the receptor. A monoclonal antibody (MoAb) of such functional properties was shown to recognize the intact intersubunit region both in the uncleaved haemagglutinin molecule and in a 25‐mer synthetic peptide comprising the intact intersubunit region. Specificity and functional studies revealed the conformation dependent recognition of the C‐terminal segment of the haemagglutinin 1 subunit by this MoAb. The binding of the antibody was shown to inhibit the trypsin‐mediated cleavage of the haemagglutinin molecule and the membrane fusion event. The enzymatic cleavage of the haemagglutinin was demonstrated to abolish antibody recognition of the infective virus suggesting an escape mechanism mediated by the functional destruction of this highly conserved region. The synthetic peptide corresponding to the intact intersubunit region is characterized by an ordered structure and is able to elicit an antibody response in BALB/c mice while its subfragments are nonimmunogenic. Furthermore, this peptide elicited a protective immune response demonstrated by in vivo experiments.

Reactions of β-oxo-α,β-bis(alkoxycarbonyl)phosphonium ylides based on P-heterocycles

The title phosphonium ylides give an ylidene oxolactone under reductive conditions and are unreactive towards aldehydes, but are degraded by hydroxylamine to form the simpler acetic ester phosphonium ylides that do react with aldehydes in the normal manner.

The effect of WSEWS pentapeptide and WSEWS-specific monoclonal antibodies on constitutive and IL-6 induced acute-phase protein production by a human hepatoma cell line, HEPG-2

Interleukin-6 receptor (IL-6R) is a member of the cytokine receptor superfamily characterised by the obligatory presence of WSXWS (Trp-Ser-X-Trp-Ser) sequence motif near the transmembrane domain. To more clearly understand the role of this motif, we treated the HepG2 hepatoma cell line with synthetic WSEWS peptide (E is glutamic acid) and checked the spontaneous and IL-6-induced production of acute-phase protein fibrinogen and C1-inhibitor (C1-INH). The peptide revealed a definitely stimulatory effect both on the constitutive synthesis of C1-INH and on the IL-6-induced fibrinogen synthesis of HepG2 cells. Monoclonal antibody specific for WSEWS pentapeptide was stimulatory for the spontaneous secretion of both fibrinogen and C1-INH. However, the IL-6-induced elevations of these acute-phase proteins were oppositely regulated, since the anti-WSEWS monoclonal antibody was inhibitory on the production of fibrinogen induced by IL-6 but strongly augmented the IL-6 induced production of C1-INH. Our study indicates that the WSEWS motif is critical in the effect of IL-6 on the acute-phase protein production influencing either the ligand binding by the WSEWS-containing receptor molecule or the signal transduction.

A SIMPLE PROOF OF THE ZEILBERGER-BRESSOUD q-DYSON THEOREM

As an application of the Combinatorial Nullstellensatz, we give a short polynomial proof of the q-analogue of Dyson’s conjecture formulated by Andrews and first proved by Zeilberger and Bressoud.

A new approach to constant term identities and Selberg-type integrals

Abstract Selberg-type integrals that can be turned into constant term identities for Laurent polynomials arise naturally in conjunction with random matrix models in statistical mechanics. Built on a recent idea of Karasev and Petrov we develop a general interpolation based method that is powerful enough to establish many such identities in a simple manner. The main consequence is the proof of a conjecture of Forrester related to the Calogero–Sutherland model. In fact we prove a more general theorem, which includes Aomotos constant term identity at the same time. We also demonstrate the relevance of the method in additive combinatorics.

Potential roles for microRNAs in inter-individual and inter-species communication

MicroRNAs are major regulators of gene expression at the posttranscriptional level. Besides being detected intracellularly, microRNAs have been found in body fluids, as well. Circulating microRNAs may have hormone like features, since they might affect distant cells as mediators of intercellular communication. MicroRNAs occurring in serum, urine, stool and saliva can be exploited as biomarkers of several diseases, and intensive research efforts are being performed in this field. MicroRNAs are also found in breast milk, and it cannot be excluded that these may act on the baby as a form of inter-individual transfer of epigenetic information. The presence of food-derived microRNAs is even more astonishing, thus plant microRNAs have been detected in the circulation, and these could be functionally active in the human/animal organism. Based on these observations, microRNAs could be involved in the transfer of gene expressional/epigenetic information between different individuals, but also between different species, even cross-kingdom. This microRNA-mediated communication might alter our concepts on the functioning of nature and on the development of diseases, as well.

On Families of Weakly Cross-intersecting Set-pairs

Let F be a family of pairs of sets. We call it an (a, b)-set system if for every set-pair (A,B) in F we have that |A| = a, |B| = b, and A ∩ B = O. Furthermore, F is weakly cross-intersecting if for any (Ai, Bi), (Aj, Bj) ∈ F with i ≠ j we have that Ai ∩ Bj and Aj ∩ Bi are not both empty. We investigate the maximum possible size of weakly cross-intersecting (a, b)-set systems. We give an explicit construction for the best known asymptotic lower bound. We introduce a fractional relaxation of the problem and prove that the best known upper bound is optimal for this case. We also provide the exact value for the case when a = b = 2.

Efficient Synthesis of Mixed Phosphonates by the Fragmentation‐Related Phosphorylation of Alcohols Applying a 2‐phosphabicyclo[2.2.2]octa‐5,7‐Diene 2‐Oxide Precursor

Abstract The photochemically induced phosphonylation of C1–C4 alcohols by the fragmentation of a P‐ethoxy 2‐phosphabicyclo[2.2.2]octa‐5,7‐diene 2‐oxide (1) yielded the phosphonates (3) with two different alkoxy group efficiently, without the slightest extent of transesterification.