Forced expression of RNF36 induces cell apoptosis

RNF36 (ring finger protein 36; alias Trif), a member of the RING zinc finger protein family, is expressed in germ cells at round spermatid stages during spermatogenesis. RING finger proteins have been implicated in a variety of functions including oncogenesis, viral replication, and developmental processes. Since no germ cell line is presently available to study the function of RNF36, in this research, we expressed RNF36 truncated and full-length proteins in COS-7 and HEK-293 cell lines to study the effect of RNF36 in somatic cells. The full-length RNF36 protein in both cell lines showed a speckled pattern in the nucleus. Truncated RNF36-1 protein with its putative nuclear localization signal (NLS) remained within the nucleus but lost the speckled pattern. The promyelocytic leukemia (PML) protein, another RING finger protein, was previously identified as present in the nucleus with a speckled pattern. Double-staining and coimmunoprecipitation analyses suggested that RNF36 colocalizes and interacts with PML. In vitro phosphorylation analysis further suggested that RNF36 nuclear localization is under the control of phosphorylation, which might be mediated by p38. Treatment with the p38 inhibitor SB203580 resulted in the cytoplasmic translocation of RNF36. Overexpression of full-length RNF36 in cells induced about half of the transfected cells to undergo cell death. The results of DNA fragmentation assays, flow cytometry assay, and TUNEL staining suggest that the death of RNF36-transfected cells was caused by apoptosis. Following further characterization of the molecular mechanism of RNF36-induced apoptosis, we found that the expression of Bax, caspase-2, and receptor-interacting protein were elevated upon RNF36 induction in test cells. These results suggest that RNF36 may interact with PML and induce cell apoptosis. We suspect that RNF36 may play a role in germ cell homeostasis during spermatogenesis.     

CD134 as target for specific drug delivery to auto-aggressive CD4<Superscript>+</Superscript>T cells in adjuvant arthritis

T cells have an important role during the development of autoimmune diseases. In adjuvant arthritis, a model for rheumatoid arthritis, we found that the percentage of CD4⁺ T cells expressing the activation marker CD134 (OX40 antigen) was elevated before disease onset. Moreover, these CD134⁺ T cells showed a specific proliferative response to the disease-associated epitope of mycobacterial heat shock protein 60, indicating that this subset contains auto-aggressive T cells. We studied the usefulness of CD134 as a molecular target for immune intervention in arthritis by using liposomes coated with a CD134-directed monoclonal antibody as a drug targeting system. Injection of anti-CD134 liposomes subcutaneously in the hind paws of pre-arthritic rats resulted in targeting of the majority of CD4⁺CD134⁺ T cells in the popliteal lymph nodes. Furthermore, we showed that anti-CD134 liposomes bound to activated T cells were not internalized. However, drug delivery by these liposomes could be established by loading anti-CD134 liposomes with the dipalmitate-derivatized cytostatic agent 5'-fluorodeoxyuridine. These liposomes specifically inhibited the proliferation of activated CD134⁺ T cells in vitro, and treatment with anti-CD134 liposomes containing 5'-fluorodeoxyuridine resulted in the amelioration of adjuvant arthritis. Thus, CD134 can be used as a marker for auto-aggressive CD4⁺ T cells early in arthritis, and specific liposomal targeting of drugs to these cells via CD134 can be employed to downregulate disease development.     

Gsh-2, a murine homeobox gene expressed in the developing brain

A novel murine dispersed homeobox gene, designated Gsh-2, is described. Analysis of cDNA sequence, including the full open reading frame, reveals an encoded homeodomain that is surprisingly similar to those of the Antennapedia-type clustered Hox genes. In addition, the encoded protein includes polyhistidine and polyalanine tracts, as observed for several other genes of developmental significance. In situ hybridizations showed Gsh-2 expression in the developing central nervous system, including the ganglionic eminences of the forebrain, the diencephalon, which gives rise to the thalamus and hypothalamus, and in the hindbrain. Furthermore, a random oligonucleotide selection and PCR amplification procedure was used to define a target DNA binding sequence, CNAATTAG, as a first step towards the identification of downstream target genes.     

SELDI-TOF mass spectrometry of High-Density Lipoprotein

Background  

High-Density Lipoprotein (HDL), one of the main plasma lipoproteins, serves as a docking station for proteins involved in inflammation, coagulation, and lipid metabolism.     

Genbank accession #: AF139098

Plant Molecular Biology -