scholarly journals The specific binding of the microtubule-associated protein 2 (MAP2) to the outer membrane of rat brain mitochondria

1989 ◽  
Vol 261 (1) ◽  
pp. 167-173 ◽  
Author(s):  
M Lindén ◽  
B D Nelson ◽  
J F Leterrier
Keyword(s):  
Rat Brain ◽  
Outer Membrane ◽  
Binding Sites ◽  
Specific Binding ◽  
Brain Mitochondria ◽  
Microtubule Associated Proteins ◽  
Cross Linkage ◽  
Associated Proteins ◽  
A Site

Purified mitochondria from rat brain contain microtubule-associated proteins (MAPs) bound to the outer membrane. Studies of binding in vitro performed with microtubules and with purified microtubule proteins showed that mitochondria preferentially interact with the high-molecular-mass MAPs (and not with Tau protein). Incubation of intact mitochondria with Taxol-stabilized microtubules resulted in the selective trapping of both MAPs 1 and 2 on mitochondria, indicating that an interaction between the two organelles occurred through a site on the arm-like projection of MAPs. Two MAP-binding sites were located on intact mitochondria. The lower-affinity MAP2-binding site (Kd = 2 x 10(-7) M) was preserved and enriched in the outer-membrane fraction, whereas the higher-affinity site (Kd = 1 x 10(-9) M) was destroyed after removing the outer membrane with digitonin. Detergent fractionation of mitochondrial outer membranes saturated with MAP2 bound in vitro showed that MAPs are associated with membrane fragments which contain the pore-forming protein (porin). MAP2 also partially prevents the solubilization of porin from outer membrane, indicating a MAP-induced change in the membrane environment of porin. These observations demonstrate the presence of specific MAP-binding sites on the outer membrane, suggesting an association between porin and the membrane domain involved in the cross-linkage between microtubules and mitochondria.

scholarly journals CLIP-170 tracks growing microtubule ends by dynamically recognizing composite EB1/tubulin-binding sites

2008 ◽  
Vol 183 (7) ◽  
pp. 1223-1233 ◽  
Author(s):  
Peter Bieling ◽  
Stefanie Kandels-Lewis ◽  
Ivo A. Telley ◽  
Juliette van Dijk ◽  
Carsten Janke ◽  
...  
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Molecular Motor ◽  
Tracking System ◽  
Family Members ◽  
Recognition Mechanism ◽  
Microtubule Associated Proteins ◽  
Specific Binding Sites ◽  
Associated Proteins

The microtubule cytoskeleton is crucial for the internal organization of eukaryotic cells. Several microtubule-associated proteins link microtubules to subcellular structures. A subclass of these proteins, the plus end–binding proteins (+TIPs), selectively binds to the growing plus ends of microtubules. Here, we reconstitute a vertebrate plus end tracking system composed of the most prominent +TIPs, end-binding protein 1 (EB1) and CLIP-170, in vitro and dissect their end-tracking mechanism. We find that EB1 autonomously recognizes specific binding sites present at growing microtubule ends. In contrast, CLIP-170 does not end-track by itself but requires EB1. CLIP-170 recognizes and turns over rapidly on composite binding sites constituted by end-accumulated EB1 and tyrosinated α-tubulin. In contrast to its fission yeast orthologue Tip1, dynamic end tracking of CLIP-170 does not require the activity of a molecular motor. Our results demonstrate evolutionary diversity of the plus end recognition mechanism of CLIP-170 family members, whereas the autonomous end-tracking mechanism of EB family members is conserved.

Melatonin-binding sites in the rat brain and pituitary mapped by in-vitro autoradiography

1989 ◽  
Vol 3 (1) ◽  
pp. 71-75 ◽  
Author(s):  
L. M. Williams
Keyword(s):  
Rat Brain ◽  
Binding Sites ◽  
Area Postrema ◽  
Specific Binding ◽  
Melatonin Receptors ◽  
Pars Tuberalis ◽  
Lower Affinity ◽  
High Affinity ◽  
In Vitro Autoradiography

ABSTRACT Using picomolar concentrations of [125I]iodomelatonin and in-vitro autoradiography, specific melatonin-binding sites have been mapped in the rat brain and pituitary. Using this same technique, high-affinity melatonin receptors had previously been identified in the suprachiasmatic nucleus (SCN) and median eminence regions of the rat hypothalamus. The presence of melatonin binding in the SCN has been confirmed, but the second area of binding has been identified as the pars tuberalis of the pituitary, and a completely novel area of binding is also reported in the area postrema. The existence of lower affinity melatonin receptors in the rat brain was also investigated using in-vitro autoradiography and higher concentrations of [125I]iodomelatonin. No further sites of specific binding were, however, disclosed.

scholarly journals Function of NF-kappa B/Rel binding sites in the major histocompatibility complex class II invariant chain promoter is dependent on cell-specific binding of different NF-kappa B/Rel subunits.

1994 ◽  
Vol 14 (5) ◽  
pp. 2926-2935 ◽  
Author(s):  
A M Brown ◽  
M W Linhoff ◽  
B Stein ◽  
K L Wright ◽  
A S Baldwin ◽  
...  
Keyword(s):  
Cell Lines ◽  
Binding Sites ◽  
Specific Binding ◽  
Regulatory Element ◽  
Invariant Chain ◽  
Nf Kappa B ◽  
Histocompatibility Complex ◽  
A Site

The promoter of the human major histocompatibility complex class II-associated invariant-chain gene (Ii) contains two NF-kappa B/Rel binding sites located at -109 to -118 (Ii kappa B-1) and -163 to -172 (Ii kappa B-2) from the transcription start site. We report here that the differential function of each of these NF-kappa B/Rel sites in several distinct cell types depends on cell-specific binding of NF-kappa B/Rel transcription factors. Ii kappa B-1 is a positive regulatory element in B-cell lines and in the Ii-expressing T-cell line, H9, but acts as a negative regulatory element in myelomonocytic and glia cell lines. In vivo protein-DNA contacts are detectable at Ii kappa B-1 in cell lines in which this site is functional as either a positive or negative regulator. Electrophoretic mobility supershift assays determine that members of the NF-kappa B/Rel family of transcription factors can bind to this site in vitro and that DNA-binding complexes that contain p50, p52, p65, and cRel correlate with positive regulation whereas the presence of p50 correlates with negative regulation. Ii kappa B-2 is a site of positive regulation in B-cell lines and a site of negative regulation in H9 T cells, myelomonocytic, and glial cell lines. In vivo occupancy of this site is observed only in the H9 T-cell line. Again, in vitro supershift studies indicate that the presence of p50, p52, p65, and cRel correlates with positive function whereas the presence of only p50 and p52 correlates with negative function. This differential binding of specific NF-kappa B/Rel subunits is likely to mediate the disparate functions of these two NF-kappa B/Rel binding sites.

Analysis of the Mechanism of Microtubule Dynamic Instability Using a UV Microbeam to Sever Elongating Microtubules

1988 ◽  
Vol 46 ◽  
pp. 122-123
Author(s):  
R.A Walker ◽  
S. Inoue ◽  
E.D. Salmon
Keyword(s):  
Dynamic Instability ◽  
Microtubule Dynamic ◽  
Gtp Hydrolysis ◽  
Abrupt Transition ◽  
Microtubule Associated Proteins ◽  
Asymmetrical Structure ◽  
Associated Proteins

Microtubules polymerized in vitro from tubulin purified free of microtubule-associated proteins exhibit dynamic instability (1,2,3). Free microtubule ends exist in persistent phases of elongation or rapid shortening with infrequent, but, abrupt transitions between these phases. The abrupt transition from elongation to rapid shortening is termed catastrophe and the abrupt transition from rapid shortening to elongation is termed rescue. A microtubule is an asymmetrical structure. The plus end grows faster than the minus end. The frequency of catastrophe of the plus end is somewhat greater than the minus end, while the frequency of rescue of the plus end in much lower than for the minus end (4).The mechanism of catastrophe is controversial, but for both the plus and minus microtubule ends, catastrophe is thought to be dependent on GTP hydrolysis. Microtubule elongation occurs by the association of tubulin-GTP subunits to the growing end. Sometime after incorporation into an elongating microtubule end, the GTP is hydrolyzed to GDP, yielding a core of tubulin-GDP capped by tubulin-GTP (“GTP-cap”).

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