Rapid treadmilling of brain microtubules free of microtubule-associated proteins in vitro and its suppression by tau

We have determined the treadmilling rate of brain microtubules (MTs) free of MT-associated proteins (MAPs) at polymer mass steady state in vitro by using [3H]GTP-exchange. We developed buffer conditions that suppressed dynamic instability behavior by ≈10-fold to minimize the contribution of dynamic instability to total tubulin-GTP exchange. The MTs treadmilled rapidly under the suppressed dynamic instability conditions, at a minimum rate of 0.2 μm/min. Thus, rapid treadmilling is an intrinsic property of MAP-free MTs. Further, we show that tau, an axonal stabilizing MAP involved in Alzheimer’s disease, strongly suppresses the treadmilling rate. These results indicate that tau’s function in axons might involve suppression of axonal MT treadmilling. We describe mathematically how treadmilling and dynamic instability are mechanistically distinct MT behaviors. Finally, we present a model that explains how small changes in the critical tubulin subunit concentration at MT minus ends, caused by intrinsic differences in rate constants or regulatory proteins, could produce large changes in the treadmilling rate.

Potential roles of osteopontin and αVβ3 integrin in the development of coronary artery restenosis after angioplasty

Angioplasty procedures are increasingly used to reestablish blood flow in blocked atherosclerotic coronary arteries. A serious complication of these procedures is reocclusion (restenosis), which occurs in 30–50% of patients. Migration of coronary artery smooth muscle cells (CASMCs) to the site of injury caused by angioplasty and subsequent proliferation are suggested mechanisms of reocclusion. Using both cultured human CASMCs and coronary atherectomy tissues, we studied the roles of osteopontin (OPN) and one of its receptors, αvβ3 integrin, in the pathogenesis of coronary restenosis. We also measured the plasma levels of OPN before and after angioplasty and determined the effect of exogenous OPN on CASMC migration, extracellular matrix invasion, and proliferation. We found that cultured CASMCs during log phase of growth and smooth muscle cell layer of the coronary atherosclerotic tissues of patients express both OPN mRNA and protein at a significantly elevated level compared with controls. Interestingly, whereas the baseline plasma OPN levels in control samples were virtually undetectable, those in patient plasma were remarkably high. We also found that interaction of OPN with αvβ3 integrin, expressed on CASMCs, causes migration, extracellular matrix invasion, and proliferation. These effects were abolished when OPN or αvβ3 integrin gene expression in CASMCs was inhibited by specific antisense S-oligonucleotide treatment or OPN-αvβ3 interaction was blocked by treatment of CASMCs with antibodies against OPN or αvβ3 integrin. Our results demonstrate that OPN and αvβ3 integrin play critical roles in regulating cellular functions deemed essential for restenosis. In addition, these results raise the possibility that transient inhibition of OPN gene expression or blocking of OPN-αvβ3 interaction may provide a therapeutic approach to preventing restenosis.

Stabilization of microtubule dynamics by estramustine by binding to a novel site in tubulin: A possible mechanistic basis for its antitumor action

The cellular targets for estramustine, an antitumor drug used in the treatment of hormone-refractory prostate cancer, are believed to be the spindle microtubules responsible for chromosome separation at mitosis. Estramustine only weakly inhibits polymerization of purified tubulin into microtubules by binding to tubulin (Kd, ≈30 μM) at a site distinct from the colchicine or the vinblastine binding sites. However, by video microscopy, we find that estramustine strongly stabilizes growing and shortening dynamics at plus ends of bovine brain microtubules devoid of microtubule-associated proteins at concentrations substantially below those required to inhibit polymerization of the microtubules. Estramustine strongly reduced the rate and extent both of shortening and growing, increased the percentage of time the microtubules spent in an attenuated state, neither growing nor shortening detectably, and reduced the overall dynamicity of the microtubules. Significantly, the combined suppressive effects of vinblastine and estramustine on the rate and extent of shortening and dynamicity were additive. Thus, like the antimitotic mechanisms of action of the antitumor drugs vinblastine and taxol, the antimitotic mechanism of action of estramustine may be due to kinetic stabilization of spindle microtubule dynamics. The results may explain the mechanistic basis for the benefit derived from combined use of estramustine with vinblastine or taxol, two other drugs that target microtubules, in the treatment of hormone-refractory prostate cancer.

Targeted ablation of the 25-hydroxyvitamin D 1α-hydroxylase enzyme: Evidence for skeletal, reproductive, and immune dysfunction

The active form of vitamin D, 1α,25-dihydroxyvitamin D [1α,25(OH)2D], is synthesized from its precursor 25 hydroxyvitamin D [25(OH)D] via the catalytic action of the 25(OH)D-1α-hydroxylase [1α(OH)ase] enzyme. Many roles in cell growth and differentiation have been attributed to 1,25(OH)2D, including a central role in calcium homeostasis and skeletal metabolism. To investigate the in vivo functions of 1,25(OH)2D and the molecular basis of its actions, we developed a mouse model deficient in 1α(OH)ase by targeted ablation of the hormone-binding and heme-binding domains of the 1α(OH)ase gene. After weaning, mice developed hypocalcemia, secondary hyperparathyroidism, retarded growth, and the skeletal abnormalities characteristic of rickets. These abnormalities are similar to those described in humans with the genetic disorder vitamin D dependent rickets type I [VDDR-I; also known as pseudovitamin D-deficiency rickets (PDDR)]. Altered non-collagenous matrix protein expression and reduced numbers of osteoclasts were also observed in bone. Female mutant mice were infertile and exhibited uterine hypoplasia and absent corpora lutea. Furthermore, histologically enlarged lymph nodes in the vicinity of the thyroid gland and a reduction in CD4- and CD8-positive peripheral T lymphocytes were observed. Alopecia, reported in vitamin D receptor (VDR)-deficient mice and in humans with VDDR-II, was not seen. The findings establish a critical role for the 1α(OH)ase enzyme in mineral and skeletal homeostasis as well as in female reproduction and also point to an important role in regulating immune function.

Letter from William Dana,

One letter requesting assistance with a claim against the Brazilian government relating to the condemned schooner Joseph Meade.

Letter from Robert A. Dobbins to William Tudor,

One letter providing detailed information on the capture of the schooner Shillelah by the Brazilian government and requesting Tudor relay to Dobbins the status of the ship.

Bill of disbursments [for the] sch[oone]r Lydia : manuscript, 1790

List of ship's expenses between Feb. 7 and May 25; dated at Aux Cayes, (Haiti) May 27, 1790; includes money paid to an interpreter at Tobago, workers for ship maintenance, and various harbor officials; and notation of purchases for goods such as sugar, coffee, rice, tar, food, and rum.

Late Miocene to Quaternary planktonic foraminifera of ODP SIte 114-704

The planktonic foraminiferal zonation of Jenkins and Srinivasan (1986), which was defined for the southwestern sector of the temperate South Pacific Ocean, was successfully extended to the temperate sequences at Site 704. The zonation is based on first and last appearances of globorotalids, principally Globoconella species, which are indigenous to temperate surface-water masses. Most of the first and last appearances at ODP Site 704 are diachronous with those in the warmer temperate to subtropical South Atlantic, North Atlantic, and South Pacific oceans. The upper Miocene, upper Pliocene, and Quaternary sequences are punctuated by frequent incursions of subantarctic and polar assemblages of planktonic foraminifers. I assume that the appearance of an assemblage dominated by sinistral Neogloboquadrina pachyderma means that the Polar Front has migrated northward, but I do not know its position north or south of the site based on this preliminary work. The upper Miocene sequence contains five incursions between 6.5 and 5 Ma and the upper Pliocene and Quaternary sequence contains 16 events since 2.47 Ma. These are minimum estimates because the number of observed events will probably increase with higher sample density and use of quantitative methods to reveal more subtle events.

Planktonic foraminiferal abundances and stable isotope ratios in middle Miocene sediments of the North Atlantic and the southwestern Pacific

A three-fold expansion of the Antarctic ice sheet at 13.60, 12.82, and 11.60 Ma has been inferred from delta18O maxima analyzed in planktonic and benthic foraminiferal tests, although accompanying changes in sea surface temperature have not been detailed. We present estimated changes in middle Miocene surface-water temperatures based on analysis of delta18O in planktonic foraminifera collected at mid-latitude Deep Sea Drilling Project sites in the North Atlantic and South Pacific oceans. We also identify periods of ice-sheet growth based on comparisons of benthic and planktonic foraminiferal delta18O values. Our results indicate: (1) a distinct cooling of the sea surface from 13.6 to 13.5 Ma immediately following a peak in ice volume at 13.6 Ma, (2) a cooling of the sea surface during a period of increasing ice volume from 13.2 to 13.0 Ma, and (3) a development of the Antarctic ice sheet during a period of cooling of the sea surface centered at 11.6 Ma.

Navigation and meteorological data during campaign TARA_20090926Z of the Tara Oceans expedition 2009-2013

The Tara Oceans Expedition (2009-2013) sampled the world oceans on board a 36 m long schooner, collecting environmental data and organisms from viruses to planktonic metazoans for later analyses using modern sequencing and state-of-the-art imaging technologies. Tara Oceans Data are particularly suited to study the genetic, morphological and functional diversity of plankton. The present dataset contains navigation and meteorological data measured during one campaign of the Tara Oceans Expedition. Latitude and Longitude were obtained from TSG data.