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E Mandelkow

Publications and source records attributed to E Mandelkow.

At least 19 recordsLinked to original sources
PubMed1992

Glycogen synthase kinase-3 and the Alzheimer-like state of microtubule-associated protein tau.

The Alzheimer-like state of tau protein includes phosphorylation by a proline-directed Ser/Thr kinase present in normal or pathological human brain. Extending earlier results on MAP kinase, we show here that the proline-directed kinase, GSK3, can induce an Alzheimer-like immune response involving several distinct and phosphorylatable epitopes at Ser-Pro motifs, as well as a gel mobility shift, similar to MAP kinase. Both kinases behave like microtubule-associated proteins in that they co-purify through cycles of assembly and disassembly, and both kinases are directly associated with paired helical filaments.

Alzheimer Disease↗
PubMed1992

Structure of striated microtubule-associated fibers of flagellar roots. Comparison of native and reconstituted states.

Several fiber systems are associated with the flagella and basal bodies of eukaryotic cells. Apart from the contractile and Ca(2+)-sensitive system-II fibers, these include the noncontractile system-I fibers that run parallel to flagellar root microtubules. Using electron microscopy and image reconstruction, we have investigated the structure of the system-I fibers of the flagellate green alga Spermatozopsis similis. The fibers were observed in three different states: (1) in situ, (2) after isolation of the intact fibers, (3) after disassembly and reconstitution of fibers in vitro from their 34 kDa subunit protein. The fibers are highly ordered; they show a constant repeat of 28 nm, they are polar, and they contain several transverse and longitudinal striations. A model is discussed showing the system-I fiber to be built from rod-like molecules with a staggered arrangement and identical polarities.

Chlorophyta↗
PubMed1992

The Alzheimer-like phosphorylation of tau protein reduces microtubule binding and involves Ser-Pro and Thr-Pro motifs.

Tau protein can be transformed into an Alzheimer-like state by phosphorylation with a kinase activity from brain [Biernat et al. (1992) EMBO J. 11, 1593-1597]. Here we show that the phosphorylation at Ser-Pro motifs strongly decreases tau's affinity for microtubules. The major reduction occurs during the first of the three main stages of phosphorylation. The data explain the lower stability of microtubules resulting from the pathological tau phosphorylation.

Alzheimer Disease↗
PubMed1992

Phosphorylation-dependent epitopes of neurofilament antibodies on tau protein and relationship with Alzheimer tau.

We have studied the phosphorylation of tau protein from Alzheimer paired helical filaments, of tau from normal human brain, and of recombinant tau isoforms. As a tool we used monoclonal antibodies against neurofilament protein [Sternberger, N., Sternberger, L. & Ulrich, J. (1985) Proc. Natl. Acad. Sci. USA 82, 4274-4276] that crossreact with tau in a phosphorylation-dependent manner. This allowed us to deduce the state of phosphorylation in normal and pathological tau, as well as antibody epitopes. The epitope of antibody SMI33 is at the first Lys-Ser-Pro sequence motif (residues 234-236) and requires an unphosphorylated Ser-235. Antibody SMI31 binds between Ser-396 (in the second Lys-Ser-Pro motif) and Ser-404, both of which must be phosphorylated. SMI34 has a conformational epitope that depends on the interaction between regions on either side of the microtubule-binding region; it also requires phosphorylation. The phosphorylatable serines detected by the SMI antibodies are part of Ser-Pro motifs and can be phosphorylated by a protein kinase activity that can be used to induce a paired helical filament-like state in human brain tau in vitro. The phosphates are incorporated in several stages that can be identified by antibody reactivity and gel shift. This suggests a role for the phosphorylation sites in Alzheimer disease, as well as the involvement of a Ser-Pro-directed protein kinase.

Alzheimer Disease↗
PubMed1992

The juvenile microtubule-associated protein MAP2c is a rod-like molecule that forms antiparallel dimers.

We have developed a procedure to isolate the microtubule-associated protein 2c (MAP2c), a juvenile form of MAP2 occurring in mammalian brain. The shape, size, self-association, and antibody interactions of MAP2c were studied. Monomeric MAP2c is an elongated molecule with a length approximately 48 nm, considerably shorter than the higher molecular weight forms MAP2a or b of adult brain. Two monoclonal antibodies whose epitopes are near the N or C terminus, respectively, are located close to the opposite ends of the MAP2c rods. This places constraints on the types of internal folding of the molecule. MAP2c self-associates into dimers and fibrous aggregates. The dimers are predominantly antiparallel and nearly in register, as judged by antibody labeling.

Animals↗
PubMed1992

The switch of tau protein to an Alzheimer-like state includes the phosphorylation of two serine-proline motifs upstream of the microtubule binding region.

The paired helical filaments (PHFs) of Alzheimer's disease consist mainly of the microtubule-associated protein tau. PHF tau differs from normal human brain tau in that it has a higher Mr and a special state of phosphorylation. However, the protein kinase(s) involved, the phosphorylation sites on tau and the resulting conformational changes are only poorly understood. Here we show that a new monoclonal antibody, AT8, records the PHF-like state of tau in vitro, and we describe a kinase activity that turns normal tau into a PHF-like state. The epitope of AT8 is around residue 200, outside the region of internal repeats and requires the phosphorylation of serines 199 and/or 202. Both of these are followed by a proline, suggesting that the kinase activity belongs to the family of proline-directed kinases. The epitope of AT8 is nearly coincident with that of another phosphorylation-dependent antibody, TAU1 [Binder, L.I., Frankfurter, A. and Rebhun, L. (1985) J. Cell Biol., 101, 1371-1378], but the two are complementary since TAU1 requires a dephosphorylated epitope.

Alzheimer Disease↗
PubMed1992

Mitogen activated protein (MAP) kinase transforms tau protein into an Alzheimer-like state.

The microtubule-associated protein tau is a major component of the paired helical filaments (PHFs) observed in Alzheimer's disease brains. The pathological tau is distinguished from normal tau by its state of phosphorylation, higher apparent M(r) and reaction with certain antibodies. However, the protein kinase(s) have not been characterized so far. Here we describe a protein kinase from brain which specifically induces the Alzheimer-like state in tau protein. The 42 kDa protein belongs to the family of mitogen activated protein kinases (MAPKs) and is activated by tyrosine phosphorylation. It is capable of phosphorylating Ser-Pro and Thr-Pro motifs in tau protein (approximately 14-16 P1 per tau molecule). By contrast, other proline directed Ser/Thr kinases such as p34(cdc2) combined with cyclin A or B have only minor effects on tau phosphorylation. We propose that MAP kinase is abnormally active in Alzheimer brain tissue, or that the corresponding phosphatases are abnormally passive, due to a breakdown of the normal regulatory mechanisms.

Alzheimer Disease↗
PubMed1992

Alzheimer-like paired helical filaments and antiparallel dimers formed from microtubule-associated protein tau in vitro.

Recent evidence from several laboratories shows that the paired helical filaments of Alzheimer's disease brains consist mainly of the protein tau in an abnormally phosphorylated form, but the mode of assembly is not understood. Here we use EM to study several constructs derived from human brain tau and expressed in Escherichia coli. All constructs or tau isoforms are rodlike molecules with a high tendency to dimerize in an antiparallel fashion, as shown by antibody labeling and chemical crosslinking. The length of the rods is largely determined by the region of internal repeats that is also responsible for microtubule binding. One unit length of the repeat domain (three or four repeats) is around 22-25 nm, comparable to the cross-section of Alzheimer PHF cores. Constructs corresponding roughly to the repeat region of tau can form synthetic paired helical filaments resembling those from Alzheimer brain tissue. A similar self-assembly occurs with the chemically cross-linked dimers. In both cases there is no need for phosphorylation of the protein.

Alzheimer Disease↗
PubMed1992

Tubulin protofilaments and kinesin-dependent motility.

Microtubules are built of tubulin subunits assembled into hollow cylinders which consist of parallel protofilaments. Thus, motor molecules interacting with a microtubule could do so either with one or several tubulin subunits. This makes it difficult to determine the structural requirements for the interaction. One way to approach the problem is to alter the surface lattice. This can be done in several ways. Proto-filaments can be exposed on their inside (C-tubules or "sheets"), they can be made antiparallel (zinc sheets), or they can be rolled up (duplex tubules). We have exploited this polymorphism to study how the motor protein kinesin attached to a glass surface interacts and moves the various tubulin assemblies. Microtubules glide over the surface along straight paths and with uniform velocities. In the case of C-tubules, approximately 40% glide similarly to microtubules, but a major fraction do not glide at all. This indicates (a) that a full cylindrical closure is not necessary for movement, and (b) that the inside surface of microtubules does not support gliding. With zinc sheets, up to 70% of the polymers move, but the movement is discontinuous, has a reduced speed, and follows along a curved path. Since zinc sheets have protofilaments alternating in orientation and polarity, this result suggests that in principle a single protofilament can produce movement, even when its neighbors cannot. Duplex microtubules do not move because they are covered with protofilaments coiled inside out, thus preventing the interaction with kinesin. The data can be explained by assuming that the outside of one protofilament represents the minimal track for kinesin, but smooth gliding requires several parallel protofilaments. Finally, we followed the motion of kinesin-coated microbeads on sea-urchin sperm flagella, from the flagellar outer doublet microtubules to the singlet microtubule tips extending from the A-tubules. No change in behavior was detected during the transition. This indicates that even if these microtubules differ in surface lattice, this does not affect the motility.

Animals↗
PubMed1991

Microtubule dynamics and microtubule caps: a time-resolved cryo-electron microscopy study.

Microtubules display the unique property of dynamic instability characterized by phase changes between growth and shrinkage, even in constant environmental conditions. The phases can be synchronized, leading to bulk oscillations of microtubules. To study the structural basis of dynamic instability we have examined growing, shrinking, and oscillating microtubules by time-resolved cryo-EM. In particular we have addressed three questions which are currently a matter of debate: (a) What is the relationship between microtubules, tubulin subunits, and tubulin oligomers in microtubule dynamics?; (b) How do microtubules shrink? By release of subunits or via oligomers?; and (c) Is there a conformational change at microtubule ends during the transitions from growth to shrinkage and vice versa? The results show that (a) oscillating microtubules coexist with a substantial fraction of oligomers, even at a maximum of microtubule assembly; (b) microtubules disassemble primarily into oligomers; and (c) the ends of growing microtubules have straight protofilaments, shrinking microtubules have protofilaments coiled inside out. This is interpreted as a transition from a tense to a relaxed conformation which could be used to perform work, as suggested by some models of poleward chromosome movement during anaphase.

Animals↗
PubMed1991

Effect of MAP2, MAP2c, and tau on kinesin-dependent microtubule motility.

By making use of DIC video microscopy to monitor microtubule motility we have studied the effect of several MAPs (MAP2, MAP2c, tau) on microtubule-kinesin interactions and microtubule gliding. Of the three MAPs tested, MAP2 interferes most strongly with kinesin-dependent microtubule motility.

Adenosine Triphosphatases↗
PubMed1979

Structural comparisons of the aggregates of tobacco mosaic virus protein.

The coat protein of tobacco mosaic virus forms numerous aggregates, including the small A-protein, the disk, and two helical forms. The structures of the disk, the helical protein forms, and the virus are compared. Most of the differences are in the conformation of the chain between residues 89 and 113, which lies in the region of protein at the center of the virus, inside the RNA. It is disordered in the disk, but has a fixed conformation in the virus and the protein helices. The differences between the virus and the two helical protein forms are largely in the conformations of arginines and carboxylic acids in this region.

Macromolecular Substances↗
PubMed1978

Paracrystalline arrays of membrane-to-membrane cross bridges associated with the inner surface of plasma membrane.

In cultured cells of the rat kangaroo PtK2 line, veils of the cell surface were observed which consisted of only plasma membrane and paracrystalline arrays of membrane-associated particles sandwiched in between. These membrane-to-membrane cross-bridging 9-to 11-nm wide particles were somewhat coumellar-shaped and were arranged on a hexagonal lattice with an interparticle distance of 16nm. At higher magnification, they revealed an unstained core, thus suggesting a ringlike substructure. Similar arrays of paracrystal-containing veils, which were rather variable in size and frequency, were also observed in other cultured cells. It is hypothesized that these paracrystals represent protein macromolecular complexes associated with the inner plasma membrane surface which crystallize when plasma membranes come into close intracellular contact and other components of the subsurface network are removed.

Animals↗