Nuclear matrix localization of high mobility group protein I(Y) in a transgenic mouse model for prostate cancer

Nuclear shape and the underlying nuclear structure, the nuclear matrix in cancer cells. Since the NM composition is considered to maintain nuclear shape and architecture, nuclear matrix proteins (NMPs) may be involved in transformation. Our laboratory has recently characterized a subset of NMPs that are associated with prostate cancer development in the transgenic adenocarcinoma of mouse prostate (TRAMP) model. One of the identified NMPs, E3E, has a similar molecular weight (22 kDa) with a protein known as HMGI(Y). HMGI(Y) belongs to a group of non-histone and chromatin-associated proteins, high-mobility-group (HMG) proteins, and it has been shown to associate with the NM. HMGI(Y) has been reported to be elevated in different types of cancer including prostate cancer. In this study, we examined the expression of HMGI(Y) protein in the NMP composition of the TRAMP model during the progression from normal to neoplasia. The expression of HMGI(Y) in the NMP extracts of three prostatic epithelial cell lines derived from a 32-week TRAMP mouse: TRAMP-C1, TRAMP-C2, and TRAMP-C3 was also examined. Using both one-dimensional and high-resolution two-dimensional immunoblot analyses, we found that: (i) HMGI(Y) is a nuclear matrix protein expressed as two protein bands with MW of 22-24 kDa and (ii) HMGI(Y) expression is correlated with neoplastic and malignant properties in late stage TRAMP prostate tumors. Overall, these findings support the evidence that HMGI(Y) can be utilized as a marker and prognostic tool for prostate cancer.     

Structure and Mechanism of Metallocarboxypeptidases

Metallocarboxpeptidases cleave C-terminal residues from peptide substrates and participate in a wide range of physiological processes, but they also contribute to human pathology. On the basis of structural information, we can distinguish between two groups of such metallopeptidases: cowrins and funnelins. Cowrins comprise protozoan, prokaryotic, and mammalian enzymes related to both neurolysin and angiotensin-converting enzyme and their catalytic domains contain 500–700 residues. They are ellipsoidal and traversed horizontally by a long, deep, narrow active-site cleft, in which the C-terminal residues are cut from oligopeptides and unstructured protein tails. The consensus cowrin structure contains a common core of 17 helices and a three-stranded β-sheet, which participates in substrate binding. This protease family is characterized by a set of spatially conserved amino acids involved in catalysis, HEXXH+EXXS/G+H+Y/R+Y. Funnelins comprise structural relatives of the archetypal bovine carboxypeptidase A1 and feature mammalian, insect and bacterial proteins with strict carboxypeptidase activity. Their ～ 300-residue catalytic domains evince a consensus central eight-stranded β-sheet flanked on either side by a total of eight helices. They also contain a characteristic set of conserved residues, HXXE+R+NR+H+Y+E, and their active-site clefts are rather shallow and lie at the bottom of a funnel-like cavity. Therefore, these enzymes act on a large variety of well-folded proteins. In both cowrins and funnelins, substrate hydrolysis follows a common general base/acid mechanism. A metal-bound solvent molecule ultimately performs the attack on the scissile peptide bond with the assistance of a strictly conserved glutamate residue.     

Structure and mechanism of metallocarboxypeptidases

Metallocarboxpeptidases cleave C-terminal residues from peptide substrates and participate in a wide range of physiological processes, but they also contribute to human pathology. On the basis of structural information, we can distinguish between two groups of such metallopeptidases: cowrins and funnelins. Cowrins comprise protozoan, prokaryotic, and mammalian enzymes related to both neurolysin and angiotensin-converting enzyme and their catalytic domains contain 500-700 residues. They are ellipsoidal and traversed horizontally by a long, deep, narrow active-site cleft, in which the C-terminal residues are cut from oligopeptides and unstructured protein tails. The consensus cowrin structure contains a common core of 17 helices and a three-stranded beta-sheet, which participates in substrate binding. This protease family is characterized by a set of spatially conserved amino acids involved in catalysis, HEXXH+EXXS/G+H+Y/R+Y. Funnelins comprise structural relatives of the archetypal bovine carboxypeptidase A1 and feature mammalian, insect and bacterial proteins with strict carboxypeptidase activity. Their approximately 300-residue catalytic domains evince a consensus central eight-stranded beta-sheet flanked on either side by a total of eight helices. They also contain a characteristic set of conserved residues, HXXE+R+NR+H+Y+E, and their active-site clefts are rather shallow and lie at the bottom of a funnel-like cavity. Therefore, these enzymes act on a large variety of well-folded proteins. In both cowrins and funnelins, substrate hydrolysis follows a common general base/acid mechanism. A metal-bound solvent molecule ultimately performs the attack on the scissile peptide bond with the assistance of a strictly conserved glutamate residue.     

Point mutations within AT-hook domains of the HMGI homologue HMGIYL1 affect binding to gene promoter but not to four-way junction DNA

High-mobility group I/Y (HMGI/Y) proteins are chromosomal proteins involved in gene and chromatin regulation. Elevated levels of HMGI/Y proteins were reported in diverse malignant tumors, and rearrangements of their genes are casually involved in the development of benign tumors. In humans, the chromosomal locus Xp22 has been often found to be affected in diverse benign mesenchymal tumors. Recent studies revealed that this region contains a retropseudogene HMGIYL1 which potentially can be activated in a way of "exonization" upon aberrations involving this region. The coding sequence of the HMGIY-L1 is highly homologous to the HMGI(Y) gene. On the protein level, both HMGIYL1 and HMGI differ at few amino acid residues, including their putative DNA-binding domains (DBDs). Here we have approached the question of whether the HMGIYL1 product would be able to adopt a role of HMGI in the context of binding to gene promoters and chromatin. Comparative binding studies, employing protein footprinting technique, revealed that HMGIYL1 has lost the ability to bind to the promoter of the interferon beta gene, but retained its high affinity for the four-way junction DNA. Our results stress the importance of particular residues within the DBDs for DNA binding and demonstrate that tight binding of HMGI/Y proteins to the four-way junction DNA can be achieved in alternative ways. The binding of HMGIYL1 to four-way junction DNA suggests that activation of the HMGIYL1 gene would yield a protein sharing some binding properties with HMG1-box proteins and histone H1. Thus, the HMGIYL1 could interplay together with these components in chromatin regulation.     

核内小体的复杂结构与多样化功能

核内小体是定位于细胞核内的无膜结构,为多蛋白-RNA复合体,通过招募相关蛋白参与基因转录、RNA剪切、表观遗传调控、肿瘤发生与抑制及抗病毒防御等多种细胞活动。明确核内小体蛋白的形成过程、功能和调控机制对研究相关疾病与病毒-宿主作用机制均具有重要意义。以下以几种核内小体蛋白为例,对核内小体的形成方式、结构与功能进行综述,并重点阐述其在抗病毒感染中的重要作用,期望为宿主抗病毒免疫机制研究提供一个新的靶标。     

Microtubule plus-end tracking proteins in differentiated mammalian cells

Differentiated mammalian cells are often characterized by highly specialized and polarized structure. Its formation and maintenance depends on cytoskeletal components, among which microtubules play an important role. The shape and dynamic properties of microtubule networks are controlled by multiple microtubule-associated factors. These include molecular motors and non-motor proteins, some of which accumulate specifically at the growing microtubule plus-ends (the so-called microtubule plus-end tracking proteins). Plus-end tracking proteins can contribute to the regulation of microtubule dynamics, mediate the cross-talk between microtubule ends, the actin cytoskeleton and the cell cortex, and participate in transport and positioning of structural and regulatory factors and membrane organelles. Malfunction of these proteins results in various human diseases including some forms of cancer, neurodevelopmental disorders and mental retardation. In this article we discuss recent data on microtubule dynamics and activities of microtubule plus-end binding proteins important for the physiology and pathology of differentiated mammalian cells such as neurons, polarized epithelia, muscle and sperm cells.