Interaction of TIP26 from a hyperthermophilic archaeon with TFB/TBP/DNA ternary complex

Interactions of TBP-interacting protein (TIP26), TBP, and TFB from a hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 with TATA-DNA were examined by electrophoretic mobility shift assay. Tk-TFB formed a ternary complex with Tk-TBP and TATA-DNA. Tk-TIP26 did not inhibit the formation of this ternary complex, but interacted with it to form a TIP26/TFB/TBP/DNA quaternary complex. This interaction is rather weak, and a large excess of Tk-TIP26 over Tk-TBP is required to fully convert the TFB/TBP/DNA ternary complex to the quaternary complex. However, determination of the concentration of Tk-TIP26 and Tk-TBP in KOD1 cells by Western blotting analysis indicated that the concentration of Tk-TIP26 is approximately ten times that of Tk-TBP, suggesting that the quaternary complex might also form in vivo.     

CHIP is a chaperone-dependent E3 ligase that ubiquitylates unfolded protein

The ubiquitin–proteasome system catalyses the immediate destruction of misfolded or impaired proteins generated in cells, but how this proteolytic machinery recognizes abnormality of cellular proteins for selective elimination remains elusive. Here, we report that the C-terminus of Hsc70-interacting protein (CHIP) with a U-box domain is an E3 ubiquitin-ligase collaborating with molecular chaperones Hsp90 and Hsc70. Thermally denatured firefly luciferase was multiubiquitylated by CHIP in the presence of E1 and E2 (Ubc4 or UbcH5c) in vitro, only when the unfolded substrate was captured by Hsp90 or Hsc70 and Hsp40. No ubiquitylating activity was detected in CHIP lacking the U-box region. CHIP efficiently ubiquitylated denatured luciferase trapped by the C-terminal region of Hsp90, which contains a CHIP binding site. CHIP also showed self-ubiquitylating activity independent of target ubiquitylation. Our results indicate that CHIP can be regarded as ‘a quality-control E3’ that selectively ubiquitylates unfolded protein(s) by collaborating with molecular chaperones.     

Distribution of Rutaceae-specific Repeated Sequences Isolated from Citrus Genomes

Three clones of dispersed repetitive sequences (MCS-26a, JA-5and JB-7) were isolated from a library of PCR products amplifiedfrom Citrus DNA using primers complementary to the minisatellitecore sequences. Distribution of these repetitive sequences inthe genomic DNA was highly variable among members of the Rutaceaefamily studied here. MCS-26a was specifically amplified in thesubfamily Aurantioideae, but not in other subfamilies of theRutaceae. Different levels of JA-5 amplification were observedamong genera in the subfamily Aurantioideae. JB-7 was widelydetected throughout the Rutaceae. These data suggest that thethree repeated sequences analysed in this study were amplifiedat different stages in the evolution of Rutaceae and that theyare useful for systematic studies of the Rutaceae. In addition,the repetitive sequences displayed a high level of restrictionfragment length polymorphism (RFLP) among Citrus species andtheir relatives, suggesting that they serve as hot spots forchanges in the genome after amplification. Copyright 2001 Annalsof Botany Company Citrus, Rutaceae, repeated sequences, DNA fingerprinting, RFLP     

Transgenic Expression of the Formin Protein Fhod3 Selectively in the Embryonic Heart: Role of Actin-Binding Activity of Fhod3 and Its Sarcomeric Localization during Myofibrillogenesis

Fhod3 is a cardiac member of the formin family proteins that play pivotal roles in actin filament assembly in various cellular contexts. The targeted deletion of mouse Fhod3 gene leads to defects in cardiogenesis, particularly during myofibrillogenesis, followed by lethality at embryonic day (E) 11.5. However, it remains largely unknown how Fhod3 functions during myofibrillogenesis. In this study, to assess the mechanism whereby Fhod3 regulates myofibrillogenesis during embryonic cardiogenesis, we generated transgenic mice expressing Fhod3 selectively in embryonic cardiomyocytes under the control of the β-myosin heavy chain (MHC) promoter. Mice expressing wild-type Fhod3 in embryonic cardiomyocytes survive to adulthood and are fertile, whereas those expressing Fhod3 (I1127A) defective in binding to actin die by E11.5 with cardiac defects. This cardiac phenotype of the Fhod3 mutant embryos is almost identical to that observed in Fhod3 null embryos, suggesting that the actin-binding activity of Fhod3 is crucial for embryonic cardiogenesis. On the other hand, the β-MHC promoter-driven expression of wild-type Fhod3 sufficiently rescues cardiac defects of Fhod3-null embryos, indicating that the Fhod3 protein expressed in a transgenic manner can function properly to achieve myofibril maturation in embryonic cardiomyocytes. Using the transgenic mice, we further examined detailed localization of Fhod3 during myofibrillogenesis in situ and found that Fhod3 localizes to the specific central region of nascent sarcomeres prior to massive rearrangement of actin filaments and remains there throughout myofibrillogenesis. Taken together, the present findings suggest that, during embryonic cardiogenesis, Fhod3 functions as the essential reorganizer of actin filaments at the central region of maturating sarcomeres via the actin-binding activity of the FH2 domain.     

A model cortical circuit for the storage of temporal sequences

Despite the fact that temporal information processing is of particular significance in biological memory systems, not much has yet been explored about how these systems manage to store temporal information involved in sequences of stimuli. A neural network model capable of learning and recalling temporal sequences is proposed, based on a neural mechanism in which the sequences are expanded into a series of periodic rectangular oscillations. Thus, the mathematical framework underlying the model, to some extent, is concerned with the Walsh function series. The oscillatory activities generated by the interplay between excitatory and inhibitory neuron pools are transmitted to another neuron pool whose role in learning and retrieval is to modify the rhythms and phases of the rectangular oscillations. Thus, a basic functional neural circuit involves three different neuron pools. The modifiability of rhythms and phases is incorporated into the model with the aim of improving the quality of the retrieval. Numerical simulations were conducted to show the characteristic features of the learning as well as the performance of the model in memory recall.     

A model of cortical memory processing based on columnar organization

Recent neurophysiological experiments using mammalian brains indicated that some cortical neurons exhibit oscillatory activities which can be of functional importance in visual perception. These findings suggest that the oscillation is an ubiquitous feature of cortical information processing carried out by columns which are receiving growing attention as functional subdivisions of cortical circuitry. On the assumption that a basic functional unit is a column comprising excitatory and inhibitory neurons, a network model of cortical memory processing which can account for these oscillations is proposed. Numerical simulations revealed that for appropriately determined parameters the network can attain memory-pattern retrieval resulting from fixed-point behaviour despite the fact that columns have the characteristic of oscillators. Received: 19 March 1993/Accepted in revised form: 23 September 1993     

A Transient Rise in Free Mg2+ Ions Released from ATP-Mg Hydrolysis Contributes to Mitotic Chromosome Condensation

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Hyperinduction of pectate lyase in Dickeya chrysanthemi EC16 by plant-derived sugars

Pectate lyase (Pel) synthesis in Dickeya chrysanthemi has been reported to be hyperinduced in planta and also in the medium containing plant extract in addition to polygalacturonate. In this study, the major components of Pel-hyperinducing fractions were found to be glucose, fructose, and sucrose by TLC and NMR. From the analysis of the sugars and their derivatives, it was found that acyclic d-hexoses with the trans relationship between C-3 and C-5 hydroxyl groups were found to be basic structure required for hyperinducing the expression of a major isozyme in infected plants (i.e. pelE). From the fact that some non-metabolizable sugars such as 2-deoxy-d-glucose and d-fucose could lead to hyperinduction and that the hyperinduction was observed only in the medium containing low concentration (<0.25%) but not higher of the sugars was added, these sugars may be considered to participate in hyperinduction as the signal rather than through their metabolism.