Protein- und RNS-Gehalt des Hypokotyls beim stationären Wachstum im Dunkeln und unter dem Einfluß von Phytochrom (Keimlinge von Sinapis alba L.)

Zusammenfassung Das Wachstum des Hypokotyls wurde an Restkeimlingen ohne Kotyledonen (Abb. 1) untersucht. Die Wachstumsgeschwindigkeit ist in dem von uns untersuchten Zeitraum sowohl im Dunkeln als auch unter dem Einfluß von P₇₃₀ (Dauer-Dunkelrot) praktisch konstant. Obgleich sich die Wachstumsgeschwindigkeiten im Dunkeln und im Dauer-Dunkelrot um den Faktor 4 unterscheiden, hat das Dunkelrot keinen signifikanten Einfluß auf den Gesamt-Proteingehalt des Hypokotyls (bzw. der durchschnittlichen Hypokotylzelle). Der Proteingehalt nimmt im Dunkeln und im Licht kontinuierlich ab. Auch der Gesamt-RNS-Gehalt zeigt innerhalb des Versuchszeitraums eine Abnahme, die unter dem Einfluß von Dunkelrot früher einsetzt als im Dunkeln. — Man kann aus den Daten der vorliegenden Arbeit schließen, daß nur ein kleiner Teil des Gesamt-Proteins und der Gesamt-RNS einer Zelle mit dem Zellwachstum unmittelbar in Verbindung gebracht werden kann.

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Protein and RNA contents of the hypocotyl during steady state growth lengthening in the dark and under the influence of phytochrome (seedlings of sinapis alba L.)

Summary Inhibition of hypocotyl lengthening by phytochrome can be regarded as a prototype of a negative photoresponse. The hypothesis has been advanced (Schopfer, 1967) that negative photoresponses are the consequence of a differential gene repression which is exerted by P₇₃₀, the active phytochrome. This hypothesis is mainly based on experiments with specific inhibitors of RNA- and protein synthesis. —The present paper is part of an experimental program which has been designed to check this hypothesis.—Continuous irradiation with standard far-red has been used to establish a virtually stationary concentration of P₇₃₀ over the whole period of experimentation (36–60 hours after sowing). To correlate more strictly the growth response of the hypocotyl with molecular changes in this organ the axis system without cotyledons has been used (Fig. 1). Even under these conditions the growth rate of the hypocotyl is nearly constant in light (continuous far-red) and dark during the whole period of experimentation (36–60 hours after sowing) (Fig. 2, 3). It is known from earlier experiments that cell division in the hypocotyl are very rare during this period and that there is virtually no increase in the DNA contents of the organ during the period of our experimentation (Weidner, 1967). Obviously the number of cells per hypocotyl is virtually constant between 36 and 60 hours after sowing. Organ (i.e. hypocotyl) lengthening is nearly exclusively due to cellular lengthening.—If we follow the protein contents of the hypocotyl we find (Fig. 4) that the total protein of the organ decreases steadily in spite of the fact that the organ grows at a constant rate. There is no significant difference in protein contents between dark-grown and far-red grown systems although the growth rates differ by a factor of 4 (Fig. 2, 3).—The situation is some-what different with respect to total RNA (Fig. 5). The RNA contents eventually decrease in far-red as well as in dark-grown systems but the decrease is significantly faster in the far-red treated systems than in the dark controls.—It is concluded that only a very small part of the total RNA and total protein of a cell can be related to the control of cellular growth. Changes in bulk RNA and bulk protein obviously do not necessarily reflect changes in the growth rate or growth capacity of an organ or a cell.

     

Adsorption of 5′-adenosine monophosphate onto precipitated calcium phosphate: Effects of inorganic polyphosphates and carbamyl phosphate

In this paper it is shown that the adsorption of 5-adenosine monophosphate (5-AMP) onto precipitated calcium phosphate exhibits a sigmoidal profile as revealed by isotherms at 45 °C. This result indicates a cooperative behavior in the adsorption of 5-AMP. The relationship between adsorption capacity and surface area of the sedimented matrix may be interpreted as an indication that there is a monolayer of the adsorbed nucleotide on the solid surface. The pH dependence of adsorption suggests that the negatively charged phosphoryl group of 5-AMP interacts with a positively charged site (possibly Ca²⁺) on the matrix surface. The adsorption of the nucleotide is markedly decreased at pH values above 8.0. The Dixon-like plot of the effect of pH suggests an inhibitory role of hydroxyl ions in the adsorption of 5-AMP. At pH 7.5, other anions such as pyrophosphate, tripolyphosphate and carbamyl phosphate also inhibit the adsorption of the nucleotide, probably by interacting with its adsorption site. We suggest that these phosphorylated molecules could have played a role in chemical evolution by modulating the amount of nucleotides adsorbed onto mineral surfaces. The significance of these phenomena in chemical evolution is discussed.     

Divalent cations modify adsorption of 5′-AMP onto precipitated calcium phosphate: A model for cation modulation of adsorptive processes in primitive aqueous environments

The adsorption of 5′-AMP onto precipitated calcium phosphate (CaPi) requires the presence of soluble calcium and this dependence exhibits a Michaelian-like behavior. This result suggests that the formation of a complex between 5′-AMP and free Ca²⁺ (CaAMP) is a prelude to the adsorption of the nucleotide in the solid matrix. At concentrations one order of magnitude higher, Mn²⁺ and Mg²⁺ can substitute for soluble Ca²⁺ in the adsorption of 5′-AMP onto solid CaPi. However, when added simultaneously with 5′-AMP to a heterogeneous mixture that contains CaPi and soluble Ca²⁺, Mn²⁺ and Mg²⁺ inhibit the adsorption of 5′-AMP in a concentration-dependent manner. This suggests the formation of complexes that are much less effective for 5′-AMP adsorption than the CaAMP complex. On the other hand, Mn²⁺ and Mg²⁺ cannot promote desorption of the nucleotide attached to the precipitate in the presence of soluble Ca²⁺ if they are added after adsorption has attained equilibrium. Although desorption of 5′-AMP can be obtained by a sequential dilution of the soluble phase with buffer and no nucleotide in a process that obeys a Langmuir equation, the lack of effect of Mn²⁺ or Mg²⁺ when adsorption has attained its maximal value suggests strong interactions between the CaAMP complex and the solid matrix when adsorption equilibrium is reached. The divalent cations present in the matrix also participate with different selectivity in the attachment of the CaAMP complex, indicating that a cation-exchange mechanism could have acted in the modulation of adsorptive/desorptive processes involving biomonomers and phosphate surfaces in primitive aqueous environments. Received: 11 December 1995 / Accepted: 5 April 1996     

Xp-duplications with and without sex reversal

Duplications in Xp including the DSS (dosage sensitive sex reversal) region cause male to female sex reversal. We investigated two patients from families with Xp duplications. The first case was one of two sisters with karyotype 46,XY, der(22), t(X;22)(p11.3;p11)mat and unambiguous female genitalia. The living sister was developmentally retarded, and showed multiple dysmorphic features and an acrocallosal syndrome. The second case was a boy with a maternally inherited direct duplication of Xp21.3-pter with the breakpoint close to the DSS locus. He had multiple abnormalities and micropenis, but otherwise unambiguous male genitalia. We performed quantitative Southern blot analysis with probes from Xp22.13 to p21.2 to define the duplicated region. Clinical, cytogenetic, and molecular data from both patients were compared with those of previously reported related cases. A comparison of the extragenital symptoms revealed no differences between patients with or without sex reversal. In both cases, the symptoms were non-specific. Among 22 patients with a duplication in Xp, nine had unambiguous female genitalia and a well-documented duplication of the DSS region. Two patients with duplication of DSS showed ambiguous external genitalia. From these data, we conclude that induction of testicular tissue may start in these patients, but that the type of genitalia depends on the degree of subsequent degeneration by a gene in DSS.     

Linkage disequilibrium analysis in Machado-Joseph disease patients of different ethnic origins

Machado-Joseph disease (MJD) is an autosomal dominant spinocerebellar degeneration originally described in families of Portuguese-Azorean ancestry. The hypothesis that its present world distribution could result from the spread of an original founder mutation has been raised. To test this possibility we have conducted a linkage disequilibrium study of markers segregating with the MJD1 locus in a total of 64 unrelated families of different geographical origins. Significant association was detected between the MJD1 locus and marker alleles at loci D14S280, D14S1050 and D14S81. All affected individuals, except one Chinese family, had allele 3 (237 bp) at D14S280. This finding is consistent with a founder effect in our MJD population. However, distinct haplotypes were observed in patients originating from the two Azorean islands showing the highest disease prevalence; therefore, the possible existence of more than one founder mutation can not be excluded with the markers currently available. Received: 27 February 1996 / Revised: 4 June 1996     

The 21-aminosteroid U-74389F increases the number of glial fibrillary acidic protein-expressing astrocytes in the spinal cord of control and wobbler mice

Summary 1. Wobbler mice suffer an autosomal recessive mutation producing severe motoneuron degeneration and dense astrogliosis, with increased levels of glial fibrillary acidic protein (GFAP) in the spinal cord and brain stem. They have been considered animal models of amyotrophic lateral sclerosis and infantile spinal muscular atrophy. 2. Using Wobbler mice and normal littermates, we investigated the effects of the membrane-active steroid Lazaroid U-74389F on the number of GFAP-expressing astrocytes and glucocorticoid receptors (GR). Lazaroids are inhibitors of oxygen radical-induced lipid peroxidation, and proved beneficial in cases of CNS injury and ischemia. 3. Four days after pellet implantation of U-74389F into Wobbler mice, hyperplasia and hypertophy of GFAP-expressing astrocytes were apparent in the spinal cord ventral and dorsal horn, areas showing already intense astrogliosis in untreated Wobbler mice. In control mice, U-74389F also produced astrocyte hyperplasia and hypertophy in the dorsal horn and hyperplasia in the ventral-lateral funiculi of the cord. 4. Givenin vivo U-74389F did not change GR in spinal cord of Wobbler or control mice, in line with the concept that it is active in membranes but does not bind to GR. Besides, U-74390F did not compete for [³H]dexamethasone binding when addedin vitro. 5. The results suggest that stimulation of proliferation and size of GFAP-expressing astrocytes by U-74389F may be a novel mechanism of action of this compound. The Wobbler mouse may be a valuable animal model for further pharmacological testing of glucocorticoid and nonglucocorticoid steroids in neurodegenerative diseases.