Kinetische Studien zur Phytochrominduzierten Proteinsynthese

Zusammenfassung In der vorliegenden Arbeit wurde geprüft, wie P₇₃₀, das aktive Phytochrom, den Proteinstoffwechsel in den Kotyledonen des Senfkeimlings (Sinapis alba L.) beeinflußt. Die Daten zur Kinetik des Proteingehalts im Dunkeln und im Dunkelrot, die Daten über die Inkorporation von Radioaktivität in die Proteinfraktion nach Applikation von ¹⁴C-Leucin (U) und die histochemischen Befunde von Häcker (1966) erlauben den Schluß, daß P₇₃₀ eine starke Proteinsynthese (Enzymund Strukturprotein) in den Kotyledonen bewirkt. Dies wird als eine Stütze für die Auffassung angesehen, daß P₇₃₀ in den Kotyledonen in erster Linie über eine differentielle Genaktivierung wirksam wird.

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Kinetical studies on phytochrome-induced protein synthesis

Summary Phytochrome control of protein metabolism has been analyzed in the cotyledons of the mustard seedling (Sinapis alba L.) which can be regarded as being representative of dicotyledonous seedlings. A mustard seedling is a closed system for nitrogen under our experimental conditions. Fig. 2 shows the kinetics of protein content in the cotyledons of the mustard seedling in darkness and under the influence of continuous far-red irradiation, i.e. under the influence of a low but virtually constant concentration of phytochrome 730. —In order to understand these two curves it must be realized that the cotyledons of the mustard seedling contain much storage protein which will be degraded during the development of the seedling. The resulting amino acids are either translocated to other parts of the seedling or used within the cotyledons for the synthesis of enzymes and structural protein. —Histochemical studies have shown (Häcker, 1966) that under the influence of far-red light the degradation of storage protein in the cotyledons is enhanced; at the same time, however, phytochrome 730 stimulates a strong de novo synthesis of structural and enzyme protein in the cotyledons. —This interpretation of the data on protein content is supported by evidence of the phytochrome-enhanced incorporation of ¹⁴C into the protein fraction after an application of ¹⁴C-Leucine (U) (Figs. 5, 6). — All present findings on phytochrome-controlled protein synthesis agree with the hypothesis that phytochrome 730 exerts its function through a differential gene activation (Mohr, 1966b).

     

Die Regulation der RNS-Synthese in Farngametophyten Durch Licht

Zusammenfassung In einer früheren Arbeit wurde gezeigt (Ohlenroth und Mohr, 1963), daß sich die Vorkeime von Dryopteris filix-mas im Blaulicht zu normalen zweidimensionalen Prothallien mit einem relativ hohen Proteingehalt entwickeln. Im Hellrot hingegen bilden sich Zellfäden (= Protonemen) aus, deren Proteingehalt bei gleicher Photosyntheseleistung wesentlich geringer ist.In der vorliegenden Arbeit wird gezeigt, daß auch der RNS-Gehalt im Blaulicht stets größer als im Hellrot ist. Die blaulichtabhängige RNS-Zunahme setzt zeitlich früher ein als die Proteinzunahme.In Umsetzexperimenten von Hellrot nach Blau manifestiert sich die morphogenetische Umsteuerung der Protonemen zeitlich eher als die blaulichtabhängige Steigerung des Proteingehalts. Kasemir und Mohr (1965) konnten zeigen, daß es sich bei dem Blaulichtprotein in erster Linie um eine Vermehrung des Strukturproteins der Chloroplasten hanhandelt. Die blaulichtabhängige RNS-Zunahme dagegen ist spätestens zum Zeitpunkt der morphologischen Umsteuerung faßbar. Dieses Ergebnis wird dadurch gestützt, daß Blaulicht im Vergleich zu Hellrot rasch einen gesteigerten Einbau von ¹⁴C, als ¹⁴C-Uridin (U) geboten, in die RNS-Fraktion verursacht. Das Blaulicht scheint in den Farnvorkeimen zwei verschiedene Vorgänge zu verursachen. 1. Steigerung einer autonomen Proteinsynthese in den Chloroplasten. 2. Auslösung oder Steigerung einer spezifischen Enzymsynthese im Cytoplasma. Die blaulichtabhängige Steigerung der RNS-Synthese scheint damit in Zusammenhang zu stehen. Die Daten der vorliegenden Arbeit werden als Indizien dafür angesehen, daß das Blaulicht seine morphogenetische Wirkung über eine differentielle Genaktivierung ausübt.

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The regulation of RNA synthesis in fern gametophytes by light

Summary Morphogenesis and metabolism of the gametophytes (= sporelings) of the common male fern Dryopteris filix-mas are controlled by visible radiation. Short wavelengths visible radiation (= blue light) leads to an increase in protein synthesis and makes possible the formation of normal two-dimensional prothallia. Under long wavelengths visible radiation (= red light) the sporelings grow as cellular filaments the protein contents of which are markedly lower than under blue irradiation even under conditions of equal rate of dry matter accumulation in red and blue (Ohlenroth and Mohr, 1963). — It is shown in the present paper that the RNA content of sporelings of the same age is always higher in blue light than in red light (Figs. 1, 3). The blue-dependent increase of RNA occurs faster than the blue-dependent increase of protein (Fig. 2). Furthermore the increase of protein per sporeling is much larger than the increase of RNA (Fig. 4). These facts are in agreement with the hypothesis that in some way or another blue light initiates differential gene activation.The blue light-dependent morphological changes which occur if we put red grown filamentous sporelings under blue light can be measured much faster than the blue light-dependent increase of the bulk protein (Figs. 5, 6). We have to conclude as we did in a previous paper (Kasemir and Mohr, 1965) that the blue light-dependent increase in the protein content of the sporelings might be mainly due to an increase of chloroplast protein. — The blue light-dependent increase of the RNA content occurs at least as fast as the morphological changes (Figs. 5, 6). This finding is supplemented by the observation (Fig. 8) that blue light markedly and rapidly stimulates the incorporation of ¹⁴C into RNA. The ¹⁴C was applied as ¹⁴C-uridine (U). — It seems that blue light causes an increase of protein synthesis in the chloroplasts as well as in the cytoplasm. Blue light-dependent RNA synthesis seems to be involved in this response. These data support the view that blue light might exert its morphogenetic control through differential gene activation.

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Die Arbeit wurde durch Sachbeihilfen der Deutschen Forschunggemeinschaft und der Stiftung Volkswagenwerk gefördert.     

The Rotary Trickle‐Bed Reactor – A New Reactor Concept for Biological Gas Purification

A new type of reactor employed to the biological gas purification is presented. The avoidance of clogging in the carrier packing is achieved by i) the use of a structured, rotating carrier packing, ii) a definite liquid irrigation regime during start‐up, operation and clean‐up time phases, iii) an on‐line determination and control of the fixed biofilm mass. A uniform biofilm thickness is generated by an optimized liquid irrigation of the carrier packing with spray nozzles. The detachment of the fixed biomass is accomplished by liquid shear forces generated with jet nozzles. The time‐scheduled operation regime of the reactor is founded on the on‐line quantification of the immobilized biomass, which results in a new quality of process governing of biotrickling reactors applied to gas purification. This is proved by the experimental results of pressure drop, dynamic liquid holdup as well as the volumetric degradation rates. The degradation of styrene was investigated in laboratory and field experiments showing a maximal volumetric degradation rate of 150 g m^–3 h^–1 at a pollutant load of 200 g m^–3 h^–1. The feasibility of this reactor prototype is demonstrated by employing it to the elimination of industrial waste gas.     

Identifying conservation hotspots in non-breeding areas: a case study of Lake Sturgeon (<Emphasis Type="Italic">Acipenser fulvescens</Emphasis>) in the Great Lakes

Limited conservation resources necessitate the prioritization of management efforts and one of the mechanisms for prioritization is the identification of hotspots. For some species of conservation concern, hotspots have been delineated, often using individual abundance as the metric. However, areas that attract individuals from multiple breeding populations may be a higher priority for conservation. These areas may have more abundant resources or habitat quality, and multiple breeding populations simultaneously benefit from conservation actions in these areas. We identified non-spawning aggregations of Lake Sturgeon Acipenser fulvescens in lakes Huron and Erie of the Great Lakes. The spawning period in Lake Sturgeon only represents a brief portion of their life and conservation goals need to include non-spawning areas. Based on 12 microsatellite loci, we determined the population of origin of individuals in the aggregations and computed a modified population-level Simpson’s diversity index (pD). Using this criterion, two hotspots were identified. The North Channel of Lake Huron had the highest diversity of represented populations (pD = 3.14). The second identified hotspot was Saginaw Bay in Lake Huron (pD = 1.78). Both hotspots had evidence of long-distance movement into the area. Monitoring of aggregations should continue to assess temporal variability in hotspot delineation.     

Frequent genes in rare diseases: panel‐based next generation sequencing to disclose causal mutations in hereditary neuropathies

Hereditary neuropathies comprise a wide variety of chronic diseases associated to more than 80 genes identified to date. We herein examined 612 index patients with either a Charcot‐Marie‐Tooth phenotype, hereditary sensory neuropathy, familial amyloid neuropathy, or small fiber neuropathy using a customized multigene panel based on the next generation sequencing technique. In 121 cases (19.8%), we identified at least one putative pathogenic mutation. Of these, 54.4% showed an autosomal dominant, 33.9% an autosomal recessive, and 11.6% an X‐linked inheritance. The most frequently affected genes were PMP22 (16.4%), GJB1 (10.7%), MPZ, and SH3TC2 (both 9.9%), and MFN2 (8.3%). We further detected likely or known pathogenic variants in HINT1, HSPB1, NEFL, PRX, IGHMBP2, NDRG1, TTR, EGR2, FIG4, GDAP1, LMNA, LRSAM1, POLG, TRPV4, AARS, BIC2, DHTKD1, FGD4, HK1, INF2, KIF5A, PDK3, REEP1, SBF1, SBF2, SCN9A, and SPTLC2 with a declining frequency. Thirty‐four novel variants were considered likely pathogenic not having previously been described in association with any disorder in the literature. In one patient, two homozygous mutations in HK1 were detected in the multigene panel, but not by whole exome sequencing. A novel missense mutation in KIF5A was considered pathogenic because of the highly compatible phenotype. In one patient, the plasma sphingolipid profile could functionally prove the pathogenicity of a mutation in SPTLC2. One pathogenic mutation in MPZ was identified after being previously missed by Sanger sequencing. We conclude that panel based next generation sequencing is a useful, time‐ and cost‐effective approach to assist clinicians in identifying the correct diagnosis and enable causative treatment considerations.

     

Match activities of elite women soccer players at different performance levels

We sought to study the physical demands and match performance of women soccer players. Nineteen top-class and 15 high-level players were individually videotaped in competitive matches, and time-motion analysis were performed. The players changed locomotor activity >1,300 times in a game corresponding to every ~4 seconds and covered 9-11 km in total. The top-class players ran 28% longer (P < 0.05) at high intensities than high-level players (1.68 +/- 0.09 and 1.33 +/- 0.10 km, respectively) and sprinted 24% longer (P < 0.05). The top-class group had a decrease (P < 0.05) of 25-57% in high intensity running in the final 15 minutes compared with the first four 15-minutes intervals, whereas the high-level group performed less (P < 0.05) high-intensity running in the last 15 minutes of each half in comparison with the 2 previous 15-minute periods in the respective half. Peak distance covered by high intensity running in a 5-minute interval was 33% longer (P < 0.05) for the top-class players than the high-level players. In the following 5 minutes immediately after the peak interval top-class players covered 17% less (P < 0.05) high-intensity running than the game average. Defenders performed fewer (P < 0.05) intervals of high-intensity running than midfielders and attackers, as well as fewer (P < 0.05) sprints than the attackers. In conclusion, for women soccer players (1) top-class international players perform more intervals of high-intensity running than elite players at a lower level, (2) fatigue develops temporarily during and towards the end of a game, and (3) defenders have lower work rates than midfielders and attackers. The difference in high-intensity running between the 2 levels demonstrates the importance of intense intermittent exercise for match performance in women soccer. Thus, these aspects should be trained intensively in women soccer.     

Neonatal rat cardiomyocytes show characteristics of nonhomotypic gap junction channels

Neonatal rat cardiomyocytes mainly coexpress the connexins Cx40, Cx43, and to a small amount Cx45, leading to potential formation of mixed (heteromeric/heterotypic) gap junction channels. Using the dual-voltage clamp technique with switching clamp circuits, the authors investigated voltage sensitivity of gap junction channels between cell pairs of Cx40, Cx43, and Cx45 stably transfected HeLa cells and compared those data to data obtained from cell pairs of cultured neonatal rat cardiomyocytes. In accordance to previously published data, the relationship between normalized conductance and transjunctional voltage (g/V(j)) was quasisymmetrical for the transfected HeLa cells, indicating homotypic gap junction channels. Boltzmann curves fitted to data obtained from neonatal rat cardiomyocyte pairs expressing both Cx40 and Cx43 showed an asymmetrical inactivation pattern, which cannot be explained by the presence of pure populations of homotypic gap junction channels of either isoform. In conclusion the authors assume the additional presence of heterotypic and possibly even heteromeric gap junction channels in neonatal rat cardiomyocytes.     

Covalent labeling of nuclear vitamin D receptor with affinity labeling reagents containing a cross-linking probe at three different positions of the parent ligand: Structural and biochemical implications

Structure-functional characterization of vitamin D receptor (VDR) requires identification of structurally distinct areas of VDR-ligand-binding domain (VDR-LBD) important for biological properties of 1α,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃). We hypothesized that covalent attachment of the ligand into VDR-LBD might alter ‘surface structure’ of that area influencing biological activity of the ligand. We compared anti-proliferative activity of three affinity alkylating derivatives of 1,25(OH)₂D₃ containing an alkylating probe at 1,3 and 11 positions. These compounds possessed high-affinity binding for VDR; and affinity labeled VDR-LBD. But, only the analog with probe at 3-position significantly altered growth in keratinocytes, compared with 1,25(OH)₂D₃. Molecular models of these analogs, docked inside VDR-LBD tentatively identified Ser237 (helix-3: 1,25(OH)₂D₃-1-BE), Cys288 (β-hairpin region: 1,25(OH)₂D₃-3-BE,) and Tyr295 (helix-6: 1,25(OH)₂D₃-11-BE,) as amino acids that are potentially modified by these reagents. Therefore, we conclude that the β-hairpin region (modified by 1,25(OH)₂D₃-3-BE) is most important for growth inhibition by 1,25(OH)₂D₃, while helices 3 and 6 are less important for such activity.