cDNA structures and regulation of two interferon-induced human Mx proteins.

Human cells treated with interferon synthesize two proteins that exhibit high homology to murine Mx1 protein, which has previously been identified as the mediator of interferon-induced cellular resistance of mouse cells against influenza viruses. Using murine Mx1 cDNA as a hybridization probe, we have isolated cDNA clones originating from two distinct human Mx genes, designated MxA and MxB. In human fibroblasts, expression of MxA and MxB is strongly induced by alpha interferon (IFN-alpha), IFN-beta, Newcastle disease virus, and, to a much lesser extent, IFN-gamma, MxA and MxB proteins have molecular masses of 76 and 73 kilodaltons, respectively, and their sequences are 63% identical. A comparison of human and mouse Mx proteins revealed that human MxA and mouse Mx2 are the most closely related proteins, showing 77% sequence identity. Near their amino termini, human and mouse Mx proteins contain a block of 53 identical amino acids and additional regions of very high sequence similarity. These conserved sequences are also present in a double-stranded RNA-inducible fish gene, which suggests that they may constitute a functionally important domain of Mx proteins. In contrast to mouse Mx1 protein, which accumulates in the nuclei of IFN-treated mouse cells, the two human Mx proteins both accumulate in the cytoplasm of IFN-treated cells.     

Prediction of "hot spots" in interleukin-2 based on informational spectrum characteristics of growth-regulating factors. Comparison with experimental data

The Resonant Recognition Model (RRM) is a theoretical method for analysis of protein and nucleotide sequences, based on the Fourier transform of the numerical representation of sequences. The amplitude spectrum of this transform is designated Informational Spectrum (IS). There are certain common frequencies in IS of growth-regulating factors. These characteristic frequencies may correlate with their roles in cell proliferation and metabolism, and in antitumor activity. IS of IL-2 has prominent characteristics in the main frequency domain of growth factors, frequency domain of antitumor factors, and frequency domain characteristic for IL-2-alpha receptor. By means of the inverse method for these 3 domains, the amino acids in the sequence of human IL-2 that may be relevant to its biological function, the so-called "hot spots", were predicted. The most probable hot spots, obtained in this way, are in the potential binding site of IL-2 to its receptor, which agrees with experimental data.     

Transgenic markers for mammalian chimeras

Summary We describe the construction of aggregation chimeras between normal and transgenic embryos containing multiple copies of mouse -globin genes. The transgenic component of the chimeras is then detected in tissue sections by a DNA-DNA in situ hybridization technique, using a biotinylated DNA -globin probe and an avidin-linked alkaline phosphatase detection system. The general advantages of transgenic markers for chimeras are discussed.     

Untersuchung der eiweißspindeln des kakteen-X-Virus mit fluoreszierenden Antikörpern

A?koliv na zá kladě mnoha pokus? se p? edpokládalo, ?e tzv. bÍlkovinná v?etena v buňkách tzn. buně?né inkluse X-viru kaktus? (Ca XV), jsou slo?ena z ?etních prodlou?ených ?ásti Ca XV, p?esto to dosud nebylo proká zá no. Proto jsme se pokusili pomocÍ fluoreskujÍcÍch protilátek doká ?at, ?e bilkovinná v?etena jsou skute?ně agregáty virových ?ástic. V těto práci jsme pouzili tzv. nep?Ímé metody. Nejprve jsme p? sobili na buňky obsahujÍci tato v?etena homologiokým antisé rem proti Ca XV, zÍskanym imunizacÍ králÍk? a teprve potom jsme buňky vlo?ili do roztoku fluoreskujicÍch protilátek proti králicimu γglobulinu. BÍlkovinná v?etena svitila potom ve fluorescen?nÍm mikroskopu silně ?lutozeleně (bylopou?ito fluoresceinisothiocyaná tu). Tato fluorescence ná m uká zala, ?e nastala pozitivnÍ reakce a ?e bÍlkovianá v?etena jsou slo?ena z virových ?ástic. ?etné kontrolnÍ pokusy potvrdily ná? základnÍ pokus.     

Resistance to influenza virus and vesicular stomatitis virus conferred by expression of human MxA protein.

MxA and MxB are interferon-induced proteins of human cells and are related to the murine protein Mx1, which confers selective resistance to influenza virus. In contrast to the nuclear murine protein Mx1, MxA and MxB are located in the cytoplasm, and their role in the interferon-induced antiviral state was unknown. In this report we show that transfected cell lines expressing MxA acquired a high degree of resistance to influenza A virus. Surprisingly, MxA also conferred resistance to vesicular stomatitis virus. Expression of MxA in transfected 3T3 cells had no effect on the multiplication of two picornaviruses, a togavirus, or herpes simplex virus type 1. Treatment of MxA-expressing cells with antibodies to mouse alpha-beta interferon did not abolish the resistance phenotype. The conclusion that resistance to influenza virus and vesicular stomatitis virus was due to the specific action of MxA is further supported by the observation that transfected 3T3 cell lines expressing the related MxB failed to acquire virus resistance.     

Identification of noncollagenous basement membrane glycopolypeptides synthesized by mouse parietal entoderm and an entodermal cell line

Using two-dimensional gel electrophoresis, we have identified two noncollagenous basement membrane (BM) glycopolypeptides which are synthesized by the mouse teratocarcinoma-derived parietal yolk sac (PYS) cell line. These glycopolypeptides have molecular weights of about 200,000 and isoelectric points of about 5.6. Polypeptides with identical parameters are synthesized by the parietal entodermal cells of mouse embryos and are found in Reichert's membrane. Pluripotent embryonal carcinoma cells (ECC) synthesize considerable amounts of the two polypeptides, whereas the yield from nullipotent ECC is negligible. The treatment of nullipotent F9 cells with retinoic acid, which induces entodermal differentiation, activates the synthesis of these polypeptides. These results indicate that the two polypeptides can be used as markers of parietal entoderm differentiation.     

The effect of phosphatidylcholine depletion on biochemical and physical properties of a Neurospora crassa membrane mutant

By using the choline starvation process it is possible to deplete the membranes of Neurospora crassa choline auxotroph $\text{chol-1}$ of phosphatidylcholine, without affecting the viability of germinated spores or whole mycelium. Spin label probes were used to examine the possible dependence of the physical state of cellular lipids on the presence of phosphatidylcholine in the membranes.Increased freedom of rotational motion of lipid soluble probes was regularly detected in choline-starved mycelium. The accumulation of neutral lipids (mostly triglycerides) in bulk form was also observed during the choline starvation process. The experiments with isolated and separated lipid classes indicated that the observed increase in fluidity of lipids in choline-starved mycelium is partly due to the difference in physical properties between bulk lipids and membrane lipids. Spin label probe 2N4 ($\text{2-}\text{propyl}\text{-2,5,5-}\text{trimethyl-oxazolidine}\text{-N-}\text{oxyl}$), which can partition at the membrane-water interface, exhibited easier partitioning among membrane lipids of choline-starved mycelium.