Binding of elements of protein kinase C-alpha regulatory domain to lamin B1

Previous results from our laboratory have demonstrated that lamin B1 is a protein kinase C (PKC)-binding protein. Here, we have identified the regions of PKC-alpha that are important for this binding. By means of overlay assays and fusion proteins made of glutathione-S-transferase (GST) fused to elements of the regulatory domain of rat PKC-alpha, we have established that binding occurs through both the V1 region and a portion of the C2 region (i.e., the calcium-dependent lipid binding [CaLB] domain) of the kinase. In particular, we have found that amino acids 200-217 of the CaLB domain are essential for binding lamin B1, as a synthetic peptide corresponding to this stretch of amino acids prevented the interaction between the CaLB domain of PKC-alpha and lamin B1. In agreement with the results of other investigators, we have determined that binding of regulatory elements of PKC-alpha to lamin B1 does not require the presence of cofactors such as PS and Ca(2+). We have also found that the binding site of lamin B1 for PKC-alpha is localized in the carboxyl-terminus of the lamin. Our findings may prove to be important in shedding more light on the mechanisms that regulate PKC functions within the nuclear compartment and may also lead to the synthesis of isozyme-specific pharmacological tools to attenuate or reverse PKC-dependent nuclear signalling pathways important for the pathogenesis of cancer.     

Endogenous growth regulator content in suspension cells of different male and female genotypes of asparagus

The endogenous levels of abscisic acid (ABA), indoleacetic acid (IAA) and of three cytokinins (transzeatin riboside, tZR; isopentenyladenosine, IPA; dihydrozeatin riboside, DHZR) were assayed in suspension cell cultures of male and female plants of asparagus by an immunological method. Assays were carried out on two different materials:

Study on the structure and properties of wool keratin regenerated from formic acid

Structural characteristics of keratin regenerated from water (KW) and from formic (KF) acid solutions were compared. Amino acid composition and molecular weight distribution of KW and KF samples were studied by high performance liquid chromatography (HPLC) and SDS-PAGE electrophoresis. Turbidity measurement showed that keratin dissolved in formic acid forms transparent and stable solutions and no flocculation occurs. In addition, because of its good solvation properties, studied by viscosity measurements, formic acid can be used as a co-solvent to prepare keratin-based blend solutions. Structural studies carried out by X-ray diffraction (XRD), Fourier transform infrared (FT-IR) and near infrared (NIR) suggest that formic acid stabilizes the beta-sheet structure. Thermogravimetric analysis (TGA) reveals a higher thermal stability of keratin regenerated from formic acid with respect to keratin regenerated from water.     

FOXP3+ T Cells Recruited to Sites of Sterile Skeletal Muscle Injury Regulate the Fate of Satellite Cells and Guide Effective Tissue Regeneration

Muscle injury induces a classical inflammatory response in which cells of the innate immune system rapidly invade the tissue. Macrophages are prominently involved in this response and required for proper healing, as they are known to be important for clearing cellular debris and supporting satellite cell differentiation. Here, we sought to assess the role of the adaptive immune system in muscle regeneration after acute damage. We show that T lymphocytes are transiently recruited into the muscle after damage and appear to exert a pro-myogenic effect on muscle repair. We observed a decrease in the cross-sectional area of regenerating myofibers after injury in Rag2^-/- γ-chain^-/- mice, as compared to WT controls, suggesting that T cell recruitment promotes muscle regeneration. Skeletal muscle infiltrating T lymphocytes were enriched in CD4⁺CD25⁺FOXP3⁺ cells. Direct exposure of muscle satellite cells to in vitro induced Treg cells effectively enhanced their expansion, and concurrently inhibited their myogenic differentiation. In vivo, the recruitment of Tregs to acutely injured muscle was limited to the time period of satellite expansion, with possibly important implications for situations in which inflammatory conditions persist, such as muscular dystrophies and inflammatory myopathies. We conclude that the adaptive immune system, in particular T regulatory cells, is critically involved in effective skeletal muscle regeneration. Thus, in addition to their well-established role as regulators of the immune/inflammatory response, T regulatory cells also regulate the activity of skeletal muscle precursor cells, and are instrumental for the proper regeneration of this tissue.     

Effect of benzyladenine on the polypeptide pattern of plastids in excised watermelon cotyledons

We have shown in a previous paper that plastids of watermelon ( Citrullus vulgaris Schrad., cv. Fairfax) cotyledons differentiate into amyloplasts when the cotyledons are grown in water and into prochloroplasts when they are grown in benzyladenine (BA) solution. In the present work we have tested whether this large difference in development of the plastids is accompanied by equally conspicuous changes in their polypeptide pattern. Cotyledons were grown for 4 days in the dark either on distilled water or on 10⁻⁵ M BA. Alternatively they were transfered to 10⁻⁵ M BA after 4 days of growth in water.
Plastids of control cotyledons had a rather simple polypeptide pattern. The only prominent protein bands were the two subunits of ribulose bisphosphate carboxylase (EC 4.1.1.39). Contamination with storage protein was present. Plastids from BA-treated cotyledons had a much more complex polypeptide pattern. No storage protein contamination was observed. Polypeptide bands present only as traces in the control and having molecular weights of 32, 44, 48 and 49 kDa increased in intensity after an exposure of only 6 h to BA. The 32-kDa band seemed to be most dependent on the presence of the hormone. Contrary to what was observed in plastids, BA seemed to have no influence on the polypeptide pattern of mitochondria.     

Incorporation and metabolism of c9,t11 and t10,c12 conjugated linoleic acid (CLA) isomers in rat brain

Conjugated linoleic acid (CLA) has been shown to exert several biological activities in different organs, in particular organs such as adipose and mammary tissue where CLA accumulates preferentially because of its high incorporation into neutral lipids. However, despite numerous studies carried out in different experimental models, both in vivo and in vitro, very little is known about the accumulation and metabolism of CLA in the brain. In this communication we present data showing that the two CLA isomers c9,t11 and t10,c12 are actively incorporated and metabolised in rat brain, and in cultures of astrocytes in vitro with patterns remarkably similar to those previously reported to occur in other tissues and cells. However, beta oxidation of CLA was found to be more efficient in brain than in other tissues, with t10,c12 a better substrate than the c9,t11 isomer. CLA incorporation and metabolism have been linked to antiinflammatory and antiproliferative activities in experimental models. Therefore, CLA activity in brain could have a positive impact on neurological disorders, such as Alzheimer's disease, Parkinson's disease and adrenoleukodystrophy, where an observed increase in inflammatory responses seems to contribute heavily to the pathogenesis.