scholarly journals Structure/Function Analysis of Xenopus Cryptochromes 1 and 2 Reveals Differential Nuclear Localization Mechanisms and Functional Domains Important forInteraction with and Repression of CLOCK-BMAL1

2007 ◽  
Vol 27 (6) ◽  
pp. 2120-2129 ◽  
Author(s):  
Ellena A. van der Schalie ◽  
Francesca E. Conte ◽  
Karla E. Marz ◽  
Carla B. Green
Keyword(s):  
Structure Function ◽  
Nuclear Localization ◽  
Molecular Clock ◽  
Feedback Loop ◽  
Cultured Cells ◽  
Function Analysis ◽  
Functional Domains ◽  
Positive Elements ◽  
A Cell

ABSTRACT Circadian rhythms control the temporal arrangement of molecular, physiological, and behavioral processes within an organism and also synchronize these processes with the external environment. A cell autonomous molecular oscillator, consisting of interlocking transcriptional/translational feedback loops, drives the approximately 24-hour duration of these rhythms. The cryptochrome protein (CRY) plays a central part in the negative feedback loop of the molecular clock by translocating to the nucleus and repressing CLOCK and BMAL1, two transcription factors that comprise the positive elements in this cycle. In order to gain insight into the inner workings of this feedback loop, we investigated the structure/function relationships of Xenopus laevis CRY1 (xCRY1) and xCRY2 in cultured cells. The C-terminal tails of both xCRY1 and xCRY2 are sufficient for their nuclear localization but achieve it by different mechanisms. Through the generation and characterization of xCRY/photolyase chimeras, we found that the second half of the photolyase homology region (PHR) of CRY is important for repression through facilitating interaction with BMAL1. Characterization of these functional domains in CRYs will help us to better understand the mechanism of the known roles of CRYs and to elucidate new intricacies of the molecular clock.

scholarly journals Regulated nuclear import of the Drosophila rel protein dorsal: structure-function analysis.

1996 ◽  
Vol 16 (3) ◽  
pp. 1103-1114 ◽  
Author(s):  
S Govind ◽  
E Drier ◽  
L H Huang ◽  
R Steward
Keyword(s):  
Amino Acids ◽  
Structure Function ◽  
Nuclear Localization ◽  
Nuclear Import ◽  
Function Analysis ◽  
Drosophila Embryo ◽  
Nuclear Targeting ◽  
Homology Region ◽  
Early Drosophila Embryo

The formation of a gradient of nuclear Dorsal protein in the early Drosophila embryo is the last step in a maternally encoded dorsal-ventral signal transduction pathway. This gradient is formed in response to a ventral signal, which leads to the dissociation of cytoplasmic Dorsal from the I kappa B homolog Cactus. Free Dorsal is then targeted to the nucleus. Dorsal is a Rel-family transcription factor. Signal-dependent nuclear localization characterizes the regulation of Rel proteins. In order to identify regions of Dorsal that are essential for its homodimerization, nuclear targeting, and interaction with Cactus, we have performed an in vivo structure-function analysis. Our results show that all these functions are carried out by regions within the conserved Rel-homology region of Dorsal. The C-terminal divergent half of Dorsal is dispensable for its selective nuclear import. A basic stretch of 6 amino acids at the C terminus of the Rel-homology region is necessary for nuclear localization. This nuclear localization signal is not required for Cactus binding. Removal of the N-terminal 40 amino acids abolished the nuclear import of Dorsal, uncovering a potentially novel function for this highly conserved region.

Characterization of phospholipase D in a cell-free system of cultured cells derived from rat frontal cortex

1992 ◽  
Vol 595 (1) ◽  
pp. 12-16 ◽  
Author(s):  
Akira Nishida ◽  
Masami Shimizu ◽  
Yasunori Kanaho ◽  
Yoshinori Nozawa ◽  
Shigeto Yamawaki
Keyword(s):  
Frontal Cortex ◽  
Phospholipase D ◽  
Cultured Cells ◽  
Free System ◽  
Cell Free System ◽  
A Cell ◽  
Rat Frontal Cortex

Structure-function analysis of the extracellular domains of the Duffy antigen/receptor for chemokines: characterization of antibody and chemokine binding sites

2003 ◽  
Vol 122 (6) ◽  
pp. 1014-1023 ◽  
Author(s):  
Christophe Tournamille ◽  
Anne Filipe ◽  
Kazimiera Wasniowska ◽  
Pierre Gane ◽  
Elwira Lisowska ◽  
...  
Keyword(s):  
Structure Function ◽  
Binding Sites ◽  
Function Analysis ◽  
Antigen Receptor ◽  
Duffy Antigen ◽  
Extracellular Domains

scholarly journals Overlapping domains of the heterochromatin-associated protein HP1 mediate nuclear localization and heterochromatin binding.

1993 ◽  
Vol 120 (2) ◽  
pp. 291-299 ◽  
Author(s):  
J A Powers ◽  
J C Eissenberg
Keyword(s):  
Amino Acids ◽  
Nuclear Localization ◽  
Functional Domains ◽  
Chromosomal Protein ◽  
Evolutionarily Conserved ◽  
Drosophila Protein ◽  
Nonhistone Chromosomal Protein ◽  
Beta Galactosidase ◽  
Drosophila Embryos

The Drosophila protein HP1 is a 206 amino acid heterochromatin-associated nonhistone chromosomal protein. Based on the characterization of HP1 to date, there are three properties intrinsic to HP1: nuclear localization, heterochromatin binding, and gene silencing. In this work, we have concentrated on the identification of domains responsible for the nuclear localization and heterochromatin binding properties of HP1. We have expressed a series of beta-galactosidase/HP1 fusion proteins in Drosophila embryos and polytene tissue and have used beta-galactosidase enzymatic activity to identify the subcellular localization of each fusion protein. We have identified two functional domains in HP1: a nuclear localization domain of amino acids 152-206 and a heterochromatin binding domain of amino acids 95-206. Both of these functional domains overlap an evolutionarily conserved COOH-terminal region.

A HOX complex, a repressor element and a 50 bp sequence confer regional specificity to a DPP-responsive enhancer

Development ◽  
2001 ◽  
Vol 128 (14) ◽  
pp. 2833-2845 ◽  
Author(s):  
Thomas Marty ◽  
M. Alessandra Vigano ◽  
Carlos Ribeiro ◽  
Ute Nussbaumer ◽  
Nicole C. Grieder ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Cultured Cells ◽  
Cell Type ◽  
Response Element ◽  
Regional Specificity ◽  
A Cell ◽  
Cell Type Specific ◽  
Hox Protein ◽  
Repressor Element

A central theme during development and homeostasis is the generation of cell type-specific responses to the action of a limited number of extant signaling cascades triggered by extracellular ligands. The molecular mechanisms by which information from such signals are integrated in responding cells in a cell-type specific manner remain poorly understood. We have undertaken a detailed characterization of an enhancer that is regulated by DPP signaling and by the homeotic protein Labial and its partners, Extradenticle and Homothorax. The expression driven by this enhancer (lab550) and numerous deletions and point mutants thereof was studied in wild-type and mutant Drosophila embryos as well as in cultured cells. We find that the lab550 enhancer is composed of two elements, a Homeotic Response Element (HOMRE) and a DPP Response Element (DPPRE) that synergize. None of these two elements can reproduce the expression of lab550, either with regard to expression level or with regard to spatial restriction. The isolated DPPRE of lab550 responds extremely weakly to DPP. Interestingly, we found that the inducibility of this DPPRE is weak because it is tuned down by the action of a repressor element. This repressor element and an additional 50 bp element appear to be crucial for the cooperation of the HOMRE and the DPPRE, and might tightly link the DPP response to the homeotic input. The cooperation between the different elements of the enhancer leads to the segmentally restricted activity of lab550 in the endoderm and provides a mechanism to create specific responses to DPP signaling with the help of a HOX protein complex.

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