Identification of AGR2 protein, a novel potential cancer marker, using proteomics technologies

A comparative analysis of the proteins in prostate tissues of the patients operated for hyperplasia (n = 7) or cancer (n = 5) was performed aiming to search for protein diagnostic markers. Differences in several minor proteins were detected using two-dimensional electrophoresis according to O'Farrel; among them, an additional protein with a molecular weight of 19 kDa and an isoelectric point of 9.0 was observed in four of the cancer cases. Mass spectrometry allowed this protein to be identified as the androgen-induced secreted protein AGR2. The possibility of using AGR2 as a diagnostic marker of prostate cancer is discussed.     

Cytogenetic analyses of eight species in the genus <Emphasis Type="Italic">Leptodactylus</Emphasis> Fitzinger, 1843 (Amphibia,Anura, Leptodactylidae), including a new diploid number and a karyotype with multiple translocations

Background

The karyotypes of Leptodactylus species usually consist of 22 bi-armed chromosomes, but morphological variations in some chromosomes and even differences in the 2n have been reported. To better understand the mechanisms responsible for these differences, eight species were analysed using classical and molecular cytogenetic techniques, including replication banding with BrdU incorporation.

Results

Distinct chromosome numbers were found: 2n = 22 in Leptodactylus chaquensis, L. labyrinthicus, L. pentadactylus, L. petersii, L. podicipinus, and L. rhodomystax; 2n = 20 in Leptodactylus sp. (aff. podicipinus); and 2n = 24 in L. marmoratus. Among the species with 2n = 22, only three had the same basic karyotype. Leptodactylus pentadactylus presented multiple translocations, L. petersii displayed chromosome morphological discrepancy, and L. podicipinus had four pairs of telocentric chromosomes. Replication banding was crucial for characterising this variability and for explaining the reduced 2n in Leptodactylus sp. (aff. podicipinus). Leptodactylus marmoratus had few chromosomes with a similar banding patterns to the 2n = 22 karyotypes. The majority of the species presented a single NOR-bearing pair, which was confirmed using Ag-impregnation and FISH with an rDNA probe. In general, the NOR-bearing chromosomes corresponded to chromosome 8, but NORs were found on chromosome 3 or 4 in some species. Leptodactylus marmoratus had NORs on chromosome pairs 6 and 8. The data from C-banding, fluorochrome staining, and FISH using the telomeric probe helped in characterising the repetitive sequences. Even though hybridisation did occur on the chromosome ends, telomere-like repetitive sequences outside of the telomere region were identified. Metaphase I cells from L. pentadactylus confirmed its complex karyotype constitution because 12 chromosomes appeared as ring-shaped chain in addition to five bivalents.

Conclusions

Species of Leptodactylus exhibited both major and minor karyotypic differences which were identified by classical and molecular cytogenetic techniques. Replication banding, which is a unique procedure that has been used to obtain longitudinal multiple band patterns in amphibian chromosomes, allowed us to outline the general mechanisms responsible for these karyotype differences. The findings also suggested that L. marmoratus, which was formerly included in the genus Adenomera, may have undergone great chromosomal repatterning.

     

Dependence of cordycepin inhibition of heterogeneous nuclear RNA biosynthesis in rat brain on the size of the poly(A) segments

A study was made of the effect of cordicepin on the biosynthesis of rat brain heterogeneous nuclear RNA (hnRNA) fractions, such as poly(A-)hnRNA, oligo(A+)hnRNA and poly(A+)hnRNA differing in the size of poly(A)-segments. Cordicepin was shown to inhibit the biosynthesis of poly(A+)hnRNA alone. However, small doses of the antibiotic do not virtually inhibit the biosynthesis of hnRNA with shorter segments of poly(A). At the same time they inhibit the biosynthesis of hnRNA containing long chains of poly(A) in the 3'-end. A possible molecular mechanism of the phenomenon reviewed is under discussion.     

Modification of recombinant asparaginase from Erwinia carotovora with polyethylene glycol 5000

A method for polyethylene conjugation with recombinant asparaginase has been developed to improve therapeutically important properties of enzyme. Methoxy-p-nitrophenyl carbamate of polyethylene glycol with molecular weight 5000 was employed as the modification reagent. Optimization of the pegylation procedure resulted in high level of enzyme modification. Under 4.5 molar excess of the modification reagent more than 10 molecules of methoxy-polyethylene bound per one asparaginase molecular. The modified asparaginase retained 57% of initial activity. A simple and efficient pegylation procedure described in this work can be used for production of asparaginase with improved therapeutic properties.     

Matrix Metalloproteinases of Normal Human Tissues

This review considers biochemical properties of the family of matrix metalloproteinases (MMPs) of normal human tissues and the involvement of these enzymes in morphogenesis. Four main MMP subfamilies are characterized, and a group of other MMPs is described. Data on mechanisms of activation and inhibition of MMPs in certain tissues during various physiological processes (embryogenesis, angiogenesis, tissue growth and involution) are considered. Information about tissue inhibitors of MMP is presented, and the ability of these inhibitors to regulate the activity of MMPs is analyzed.     

Modification of the half-life of poly (A+)mRNA in liver and brain cells of the rat during embryogenesis and postnatal development

The poly(A+)/poly(A-)mRNA ratio and the half-life time of poly(A+)mRNA for mRNA metabolism in the liver and brain of rat in the course of ontogensis, late embryogenesis, postnatal development and upon ageing were determined. It was shown that in the course of ontogenesis both the ratio of poly(A+)/poly(A-)mRNA of free and membrane-bound polyribosomes and the half-life time of poly(A+)mRNA determined from the degradation kinetics in the presence of actinomycin D are changed. A possible role of poly(A) sequences in the regulation of mRNA life-time is discussed.     

Role of matrix metalloproteinases in development of diabetic nephropathy

This review considers molecular mechanisms that underlie disorders in the structure and metabolism of renal extracellular matrix in diabetic nephropathy. The contribution of the increased synthesis of renal extracellular matrix proteins in the accumulation of renal mesangial matrix is considered, and the important role of the degradation system of the extracellular matrix proteins in the development of fibrosis is also shown. Data on changes in mRNA expression for the matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) in various forms of diabetic nephropathy are presented. A correlation is established between changes in the balance of MMP proteolytic activity and TIMP activity and the accumulation of extracellular matrix.     

Identification of AGR2 protein, a novel potential cancer marker, using proteomics technologies

A comparative analysis of the proteins in prostate tissues of the patients operated for hyperplasia (n = 7) or cancer (n = 5) was performed aiming to search for protein diagnostic markers. Differences in several minor proteins were detected using two-dimensional electrophoresis according to O’Farrel; among them, an additional protein with a molecular weight of 19 kDa and an isoelectric point of 9.0 was observed in four of the cancer cases. Mass spectrometry allowed this protein to be identified as the androgen-induced secreted protein AGR2. The possibility of using AGR2 as a diagnostic marker of prostate cancer is discussed.     

Role of matrix metalloproteinases and their inhibitors in tumor invasion and metastasis

The role of various matrix metalloproteinases (MMP)—such as gelatinases, stromelysins, matrilysin, collagenase-3, and membrane-bound MMP (MB-MMP)—in tumor invasion and metastasis is discussed. Data suggesting significance for malignant growth of the expression level of these enzymes and also of their activators and inhibitors are presented. It is concluded that at different stages of tumor progression the activity of different MMPs is displayed, which is regulated by various growth factors and oncogenes. Different malignancies are characterized by changes in activities of specific MMPs. Data are presented which show significance of the ratio between the MMP activity and that of tissue inhibitors of metalloproteinases (TIMP) in tumor invasion and metastasis, especially in connection with a dual role of TIMP as both MMP inhibitors and activators.