SUMOylation and PARylation cooperate to recruit and stabilize SLX4 at DNA damage sites

SUMOylation plays important roles in the DNA damage response. However, whether it is important for interstrand crosslink repair remains unknown. We report that the SLX4 nuclease scaffold protein is regulated by SUMOylation. We have identified three SUMO interaction motifs (SIMs) in SLX4, mutating all of which abrogated the binding of SLX4 to SUMO-2 and covalent SLX4 SUMOylation. An SLX4 mutant lacking functional SIMs is not recruited to PML nuclear bodies nor stabilized at laser-induced DNA damage sites. Additionally, we elucidated a novel role for PARylation in the recruitment of SLX4 to sites of DNA damage. Combined, our results uncover how SLX4 is regulated by post-translational modifications.     

Activating and inhibitory Fcγ receptors in rheumatoid arthritis: from treatment to targeted therapies

Fcγ receptors (FcγRs) bind the constant Fc region of IgG molecules. IgG/antigen-containing immune complexes elicit a variety of effector functions in cells that express activating FcγRs. Because activating FcγRs are present on cells from the innate immune system, such as dendritic cells, monocytes/macrophages and granulocytes, these IgG receptors form a crucial link between the innate and the acquired immune systems. Recently, the ability to detect the inhibitory FcγRIIb on cells has indicated an imbalance between activating and inhibitory FcγRs in rheumatoid arthritis. This progress offers an opportunity to study modulation of FcγR balance and could stimulate development of FcγR-directed immunotherapy.     

Effect of diiodobenzotefa on the functionally different subpopulations of T-lymphocytes]

Diiodobenzo-tepa (DIB) was given orally to CBA mice in a dose of 25 mg/kg for 3 successive days. The number of nucleus-containing cells decreased 3.9 fold in the thymus and 1.4 fold in the bone marrow. In experiments on transplantation of lymphoid cells to intact or lethally irradiated (CBA X C57BL/6J)F1 mice treated with DIB this substance did not influence the helper activity of T lymphocytes but inhibited the activity or B and T lymphocytes, inducing "graft-versus-host" and T cell-suppressor functions.     

Length variation and secondary structure of introns in the Mlc1 gene in six species of Drosophila

A nearly universal feature of intron sequences is that even closely related species exhibit a large number of insertion/deletion differences. The goal of the analysis described here is to test whether the observed pattern of insertion/deletion events in the genealogy of the myosin alkali light chain (Mlc1) gene is consistent with neutrality, and if not, to determine the underlying forces of evolutionary change. Mlc1 pre-mRNA is alternatively spliced, and one constraint is that signals necessary for tissue-specificity of directed splicing must be conserved. If the total length of an intron is functionally constrained, then the distribution of indels on branches of the gene genealogy should reflect a departure from randomness. Here we perform a phylogenetic analysis, inferring ancestral states wherever possible on a phylogeny of 29 alleles of Mlc1 from six species of Drosophila. Observed patterns of indels on the genealogy were compared to those from simulated data, with the result that we cannot reject the null hypothesis of neutrality. A clear departure from a neutral prediction was seen in the excess folding free energy predicted for the introns flanking the alternatively spliced exon. Relative rate tests also suggest a retardation in the rate of Mlc1 sequence evolution in the simulans clade.      

Conservation of alternative splicing and genomic organization of the myosin alkali light-chain (Mlc1) gene among Drosophila species

The Mlc1 gene of Drosophila melanogaster encodes two MLC1 isoforms via developmentally regulated alternative pre-mRNA splicing. In larval muscle and tubular and abdominal muscles of adults, all of the six exons are included in the spliced mRNA, whereas, in the fibrillar indirect flight muscle of adult, exon 5 is excluded from the mRNA. We show that this tissue-specific pattern of alternative splicing of the Mlc1 pre-mRNA is conserved in D. simulans, D. pseudoobscura, and D. virilis. Isolation and sequencing of the Mlc1 genes from these three other Drosophila species have revealed that the overall organization of the genes is identical and that the genes have maintained a very high level of sequence identity within the coding region. Pairwise amino acid identities are 94%-99%, and there are no charge changes among the proteins. Total nucleotide divergence within the coding region of the four genes supports the accepted genealogy of these species, but the data indicate a significantly higher rate of amino acid replacement in the branch leading to D. pseudoobscura. A comparison of nucleotide substitutions in the coding portions of exon 5 and exon 6, which encode the alternative carboxyl termini of the two MLC1 isoforms, suggests that exon 5 is subject to greater evolutionary constraints than is exon 6. In addition to the coding sequences, there is significant sequence conservation within the 5' and 3' noncoding DNA and two of the introns, including one that flanks exon 5. These regions are candidates for cis- regulatory elements. Our results suggest that evolutionary constraints are acting on both the coding and noncoding sequences of the Mlc1 gene to maintain proper expression and function of the two MLC1 polypeptides.      

Role of interleukin-7 in degenerative and inflammatory joint diseases

IL-7 is known foremost for its immunostimulatory capacities, including potent T cell-dependent catabolic effects on bone. In joint diseases like rheumatoid arthritis and osteoarthritis, IL-7, via immune activation, can induce joint destruction. Now it has been demonstrated that increased IL-7 levels are produced by human articular chondrocytes of older individuals and osteoarthritis patients. IL-7 stimulates production of proteases by IL-7 receptor-expressing chondrocytes and enhances cartilage matrix degradation. This indicates that IL-7, indirectly via immune activation, but also by a direct action on cartilage, contributes to joint destruction in rheumatic diseases.     

Antiviral Activity of Trappin-2 and Elafin In Vitro and In Vivo against Genital Herpes

Serine protease inhibitor elafin (E) and its precursor, trappin-2 (Tr), have been associated with mucosal resistance to HIV-1 infection. We recently showed that Tr/E are among principal anti-HIV-1 molecules in cervicovaginal lavage (CVL) fluid, that E is ∼130 times more potent than Tr against HIV-1, and that Tr/E inhibited HIV-1 attachment and transcytosis across human genital epithelial cells (ECs). Since herpes simplex virus 2 (HSV-2) is a major sexually transmitted infection and risk factor for HIV-1 infection and transmission, we assessed Tr/E contribution to defense against HSV-2. Our in vitro studies demonstrated that pretreatment of endometrial (HEC-1A) and endocervical (End1/E6E7) ECs with human Tr-expressing adenovirus (Ad/Tr) or recombinant Tr/E proteins before or after HSV-2 infection resulted in significantly reduced virus titers compared to those of controls. Interestingly, E was ∼7 times more potent against HSV-2 infection than Tr. Conversely, knockdown of endogenous Tr/E by small interfering RNA (siRNA) significantly increased HSV-2 replication in genital ECs. Recombinant Tr and E reduced viral attachment to genital ECs by acting indirectly on cells. Further, lower viral replication was associated with reduced secretion of proinflammatory interleukin 8 (IL-8) and tumor necrosis factor alpha (TNF-α) and decreased NF-κB nuclear translocation. Additionally, protected Ad/Tr-treated ECs demonstrated enhanced interferon regulatory factor 3 (IRF3) nuclear translocation and increased antiviral IFN-β in response to HSV-2. Lastly, in vivo studies of intravaginal HSV-2 infection in Tr-transgenic mice (Etg) showed that despite similar virus replication in the genital tract, Etg mice had reduced viral load and TNF-α in the central nervous system compared to controls. Collectively, this is the first experimental evidence highlighting anti-HSV-2 activity of Tr/E in female genital mucosa.