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J Horwitz

Publications and source records attributed to J Horwitz.

At least 145 records · Page 8Linked to original sources
PubMed1985

Partial characterization of three distinct populations of human gamma-crystallins.

Certain low molecular weight crystallins may be involved preferentially in the process of human cataractogenesis. Three distinct populations of monomeric crystallins with molecular weights ranging from 19,000 to 24,000 have been demonstrated previously by electrophoresis in the presence of sodium dodecyl sulfate (SDS). The authors now report the chromatographic separation of these three components from young human lenses. Each of the three species has unique profiles on isoelectric focusing and native polyacrylamide gels as well as on SDS gels. All three species show similar patterns of age-related modification in their charge characteristics. These three populations of polypeptides all appear to be gamma-crystallins. The relative amounts of the three vary with age, which accounts for the greater ease of their separation in extracts from young lenses as compared with adult lenses.

Aged↗
PubMed1984

High molecular weight aggregates from human cataracts: characterization by Western blot analysis.

The formation of covalently linked, high molecular weight protein aggregates has been thought to play an important role in opacification of the human lens. Antisera were used in Western blot analysis to demonstrate the involvement of all major classes of lens proteins (alpha, beta and gamma crystallin; the major intrinsic membrane polypeptide) in covalent aggregation. Of these classes, aggregation of gamma and beta crystallins via intermolecular disulfide bonding and aggregation of the major intrinsic membrane polypeptide via intermolecular, non-disulfide bonding were more pronounced in cataractous as compared with normal lenses.

Cataract↗
PubMed1984

Dietary restriction retards age-related loss of gamma crystallins in the mouse lens.

The soluble crystallins in lenses from diet-restricted and control mice of diverse ages (2, 11, or 30 months) were studied by high-performance liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results obtained with both methods suggest that dietary restriction decelerates age-related loss of soluble gamma crystallins.

Aging↗
PubMed1984

The major intrinsic protein (MIP) of the bovine lens fiber membrane: characterization and structure based on cDNA cloning.

Synthetic oligonucleotide probes have been used to identify two overlapping cDNA clones that represent the entire coding region of the mRNA for the major intrinsic protein (MIP) of bovine lens cell membrane. Hybridization studies indicate that bovine MIP is encoded by a single-copy gene. The cDNA hybridizes to the rat genome, but MIP mRNA is not detected in rat liver. Analysis of the deduced amino acid sequence provides support for the potential role of MIP as a junctional protein. The structure predicted for MIP suggests that it traverses the lipid bilayer six times with both carboxy and amino termini on the cytoplasmic side, and that it has at least one amphiphilic transmembrane segment, as expected if the protein were to participate in the formation of an aqueous channel.

Amino Acid Sequence↗
PubMed1984

Clinicobiochemical correlations in aging-related human cataract. The Pan American Association and American Journal of Ophthalmology lecture.

In a series of 50 consecutive adult patients (24 men and 26 women) we studied aging-related cataract by means of comprehensive presurgical examination of the patient, the eyes, and the cataractous lens; cataract sample collection from predetermined sites during surgery; biochemical analyses with high-performance liquid chromatography of each individual lens sample; and computer-assisted biostatistical data analysis. Compared with normal values established on clear human lenses obtained from a donor eye program, the ratios of 23,000 (23K) crystallin to total beta, gamma, and low molecular weight soluble proteins were statistically significantly lower in the anterior axial lens cortex (P less than .001) and in the lens nucleus (P less than .001). Increased grades of cortex opacification and nucleus opacification were generally associated with progressively decreased 23K crystallin ratios. In this series of patients with diverse cataract-associated systemic diseases, family histories, ophthalmic disorders, and drug-use patterns, decreased 23K crystallin ratios were a sensitive marker for aging-related cataract. Similar biochemical abnormalities in patients with multiple cataract-associated factors may have important clinical implications.

Adult↗
PubMed1984

Activation of tyrosine hydroxylase in the superior cervical ganglion by nicotinic and muscarinic agonists.

Both dimethylphenylpiperazinium (DMPP), a nicotinic agonist, and bethanechol, a muscarinic agonist, increase 3,4-dihydroxyphenylalanine (DOPA) synthesis in the superior cervical ganglion of the rat. DMPP causes approximately a fivefold increase in DOPA accumulation in intact ganglia whereas bethanechol causes about a two-fold increase in DOPA accumulation. These effects are additive with each other and with the increase in DOPA accumulation produced by 8-bromo cyclic AMP. The action of DMPP is dependent on extracellular Ca2+ while the actions of bethanechol and 8-bromo cyclic AMP are not dependent on extracellular Ca2+. Cholinergic agonists and cyclic nucleotides produce a stable activation of tyrosine hydroxylase (TH) in the ganglion. The activation of TH by nicotinic and muscarinic agonists can be detected after 5 min of incubation of the ganglia with these agents. The nicotinic response disappears after 30 min of incubation, whereas the muscarinic response persists for at least 30 min. The Ca2+ dependence of the TH activation produced by these agents is similar to the Ca2+ dependence of their effects on DOPA accumulation in intact ganglia. These data are consistent with the hypothesis that nicotinic agonists, muscarinic agonists, and cyclic AMP analogues increase TH activity by three distinct mechanisms. The activation of TH presumably underlies the increase in DOPA synthesis produced by these agents.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1984

Stimulation of DOPA synthesis in the superior cervical ganglion by veratridine.

We have investigated the effect of veratridine on DOPA (3,4-dihydroxyphenylalanine) accumulation by the superior cervical ganglion of the rat. Incubation of the ganglion with veratridine (50 microM) causes a 10-fold increase in the rate of DOPA accumulation. Veratridine-stimulated DOPA accumulation is blocked by tetrodotoxin, but not by cholinergic or adrenergic antagonists or by decentralization of the ganglion. The cyclic nucleotide 8-bromo cyclic GMP does not increase DOPA accumulation, and 8-bromo cyclic AMP causes only a 2-fold increase in DOPA accumulation, which is additive with the effect of veratridine. Thus, the action of veratridine appears to be independent of these cyclic nucleotides. The effect of veratridine on DOPA accumulation is probably due to a stable modification of tyrosine hydroxylase, since an increase in tyrosine hydroxylase activity can be measured in cell-free extracts of veratridine-treated ganglia. Both the increase in DOPA accumulation and the stable activation of tyrosine hydroxylase are dependent upon extracellular Ca2+. The activation of tyrosine hydroxylase by veratridine may be mediated by the depolarization of, and the subsequent entry of Ca2+ into, ganglionic neurons.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1984

Cloning and characterization of a cow beta crystallin cDNA.

We have isolated and sequenced a beta crystallin cDNA clone derived from the mRNA of cow lenses. Comparisons of the deduced amino acid sequence with the amino acid sequences of the principal beta crystallins of the cow, Beta Bp (1) and the mouse (2,3), confirm the general homology within the beta crystallin family and with the gamma crystallins. This beta crystallin cDNA, designated pBeta25/23, has 96% amino acid homology with the murine beta23 and only 43% amino acid homology with the cow beta Bp. The N-terminal 14 amino acids of the murine beta23, which are composed of a high percentage of hydrophobic amino acid residues, bear no similarity to the predicted amino acid sequence of the cow beta crystallin cDNA clone which we have isolated. The remaining amino acid sequences show greater homology between the mouse and cow beta crystallins than the corresponding alpha and beta globins between the two species. The degree of homology is comparable to that of the alpha crystallins for the cow and mouse. The pBeta25/23 cDNA is interesting for the presence of two potential translation initiation sites which are in phase and, if both are used, would code for two polypeptides of MW 25,100 and 23,200. These correspond to the 25,000 and 23,000 dalton beta crystallins that have been partially sequenced by Berbers et al. (4). This raises the possibility that the beta 25 and the beta23 synthesized in the cow lens are not encoded by two separate genes but may be derived from single mRNA using two translation initiation sites or by post-translational processing of the larger peptide. It is possible also that the two proteins are from two mRNA's derived from a single hnRNA by differential splicing of a intervening sequence at the 5' end of the gene.

Amino Acid Sequence↗
PubMed1984

Muscarine increases tyrosine 3-monooxygenase activity and phospholipid metabolism in the superior cervical ganglion of the rat.

Muscarinic agonists cause a stable activation of tyrosine 3-monooxygenase in the superior cervical ganglion and increase the incorporation of 32Pi into phospholipids in the ganglion. We have studied the relationship between muscarine-stimulated phospholipid turnover and the muscarine-induced activation of tyrosine 3-monooxygenase. Both effects of muscarine are apparent within 2 min of incubation, and both are essentially independent of extracellular Ca++. All concentrations of muscarine that increase dopa synthesis also stimulate phospholipid turnover. Bethanechol is less efficacious than muscarine in producing both of these effects. Lithium, which disrupts phospholipid metabolism, inhibits the muscarine-stimulated accumulation of dopa. Other agents which affect phospholipid metabolism, including phospholipase C and deoxycholate, also increase the synthesis of dopa in the ganglion. These data support the hypothesis that changes in phospholipid metabolism mediate the activation of tyrosine 3-monooxygenase by muscarinic agonists.

Animals↗
PubMed1983

Protein carboxyl methyltransferase from cow eye lens.

Protein carboxyl methyltransferase activity (S-adenosyl-L-methionine: protein carboxyl-0-methyltransferase; E.C. 2.1.1.24) has been detected in crude soluble extracts of cow eye lens. The activity incorporates methyl groups from S-adenosyl-L-methionine into endogenous lens proteins in vitro, and several of these species co-migrate electrophoretically with lens crystallins. A 2600-fold purification of the enzyme free of endogenous substrates was achieved by gel filtration and affinity chromatography. The lens methyltransferase has a native molecular weight of approximately 27,000, and catalyzes the substoichiometric incorporation of highly alkali-labile methyl ester groups into a broad range of protein substrates. The lens enzyme appears to be similar to that found in human erythrocytes, which specifically recognizes and modifies D-aspartic acid residues in aged proteins in a postulated degradative or racemization-repair pathway (McFadden, P.N., and Clarke, S. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 2460-2464).

Animals↗
PubMed1983

Major intrinsic polypeptide of lens membrane. Biochemical and immunological characterization of the major cyanogen bromide fragment.

A protein of Mr 26000 has been shown to be the major component of eye-lens junctions, which are similar but not identical to the gap junctions of liver and other tissues. Cyanogen bromide cleavage of the Mr 26000 polypeptide from bovine lenses yields a major fragment of Mr 15000 (fragment 1). However, if the junctions are first treated with trypsin or carboxypeptidase Y, cyanogen bromide treatment yields a fragment of reduced molecular weight. Since protease treatment has been shown to cleave residues almost exclusively from the carboxy-terminal end of the Mr 26000 polypeptide, it follows that fragment 1 represents the carboxy-terminal half of this molecule, part of which is exposed to proteolytic attack outside the membrane. This latter result is corroborated by the fact that antisera which recognize both the Mr 26000 polypeptide and fragment 1 fail to do so after preadsorption with intact membranes. In addition, comparative amino acid and partial sequence analyses of the Mr 26000 polypeptide and fragment 1 indicate that fragment 1 is more hydrophilic in character, suggesting that much of the amino-terminal half of the Mr 26000 polypeptide is buried within the lipid bilayer.

Amino Acid Sequence↗
PubMed1983

Presence of low molecular weight polypeptides in human brunescent cataracts.

Microdissected sections from opaque & brunescent lens nuclei contain low molecular weight (4,000-8,000 dalton) polypeptides not found in microdissected sections of transparent lens nuclei. Tryptic digestion of these polypeptides from different cataracts reveal similar peptide maps. Together, these results support the involvement of proteolysis in human cataractogenesis, and suggest the possibility of similar molecular mechanisms occurring in cleavage of lens polypeptides during formation of the opaque & brunescent human cataract.

Aged↗
PubMed1983

Immunocytochemical localization of the main intrinsic polypeptide (MIP) in ultrathin frozen sections of rat lens.

The in situ distribution of the 26-kdalton Main Intrinsic Polypeptide (MIP or MP 26), a putative gap junction protein in ocular lens fibers, was defined at the electron microscope level using indirect immunoferritin labeling of ultrathin frozen sections of rat lens. MIP was found distributed throughout the plasma membrane of the lens fiber cell, with no apparent distinction between junctional and nonjunctional membrane. MIP was not detectable in the basal or lateral plasma membrane of the lens epithelial cell, including the interepithelial cell gap junctions; nor was MIP detectable in the plasma membrane or gap junctions of the hepatocyte. Previous reports have indicated that the protein composition of the lens fiber cell junction differs from that of the hepatocyte gap junction. The evidence presented here suggests that the composition of the fiber cell junction and plasma membrane is also immunocytochemically distinct from that of its progenitor, the lens epithelial cell.

Animals↗
PubMed1982

The effect of development of thermoregulatory function on the biochemical assessment of the ontogeny of neonatal dopaminergic neuronal activity.

Previous studies from this laboratory on the functional ontogeny of impulse flow in the nigrostriatal pathway of the neonatal rat were interpreted as indicating that there was an abrupt onset of impulse flow between 6 and 8 days of age. The functional development was assessed by determining the age at which the biochemical responses of these neurons to physical or pharmacological manipulation were similar to those of adult neurons. Further studies of these phenomena demonstrate that the age-dependency of these biochemical markers of impulse traffic are directly related to the development of thermoregulatory function in the neonatal rat. Accordingly, provided that 4-day-old or even younger rats are maintained at or near litter temperature, they exhibit qualitatively all of the biochemical responses characteristic of the 10-day-old or adult animal in which impulse flow is known to be present.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1982

Immunocytochemical localization of the lens main intrinsic polypeptide (MIP26) in communicating junctions.

Plasma membranes of vertebrate lens fiber cells contain a major intrinsic polypeptide with an apparent molecular weight of 26,000 (MIP26). These plasma membranes are extremely rich in communicating junctions, and it has been suggested that MIP26 is a component of them. MIP26 was purified from cow lenses using preparative SDS gel electrophoresis followed by hydroxylapatite column chromatography. From gel electrophoresis patterns and aggregational properties it was concluded that the MIP26 preparation was homogeneous. The purified MIP26 was used to produce monospecific antibodies in rabbits as assessed by double immunodiffusion and crossed immunoelectrophoresis of purified MIP26 and solubilized lens plasma membranes against the antiserum. Indirect immunocytochemical studies were performed on open and closed lens plasma membrane vesicles by incubation in anti-MIP antiserum followed by ferritin-conjugated goat antirabbit IgG. The conjugate bound unequivocally to lens communicating junctions, indicating that MIP26 is a component of these structures.

Aquaporins↗