An mRNA that specifically accumulates in maize roots delineates a novel subset of developing cortical cells

A near full-length cDNA clone (pZRP3) corresponding to an mRNA that accumulates specifically in roots of maize was isolated. The ZRP3 mRNA is ca. 600 nucleotides in length. The amino acid sequence of the predicted polypeptide is rich in leucine (16%), proline (11%), and cysteine (8.5%). The zrp3 gene appears to be expressed exclusively in roots, whereas other ZRP3-related genes are expressed in additional organs of the maize plant. In situ hybridization shows that ZRP3 mRNA accumulation is largely confined to the cells of the cortical ground meristem. Furthermore, accumulation of this mRNA occurs within a distinct subset of cortical cells, the inner three to four cell layers.Journal paper number J-14572 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa Project Number 2997.     

Regulation of stem elongation in chinese cabbage by inflorescence removal and application of growth regulators

The effect of inflorescence removal on stem elongation in Chinese cabbage cv. Spring A was studied. Removal of the inflorescence before its visibility, or upon its appearance but before the beginning of bolting (stages 1–3), markedly reduced the stem length. Removal after the beginning of bolting (stage 5) had no effect on stem length.Application of GA₃ to the treated plants partially or fully restored the elongation of the flowering stem, whereas paclobutrazol inhibited the elongation of the treated, as well as the control stems. Indole-3-acetic acid (IAA) or kinetin was ineffective in restoring stem elongation of the plants from which the inflorescence had been removed. Inflorescences at stages 1–2 were found to secrete about 10 times more gibberellic acid (GA)-like activity compared with control apices or inflorescences at stage 5.It is suggested that the developing inflorescence is the major source of GAs which control stem elongation. However, shortly after the appearance of the inflorescence at the onset of bolting, stem elongation is no longer dependent on GAs derived from the apical inflorescence but require GAs from other sources.Contribution from the Agricultural Research Organization, The Volcani Center Bet Dagan, Israel No. 2218-E, 1987 series.     

Localization of anti-clathrin antibody in the sarcomere and sensitivity of myofibril structure to chloroquine suggest a role for clathrin in myofibril assembly

Immunofluorescence microscopy has been used to demonstrate that X22, a monoclonal antibody specific for clathrin heavy chain, localizes in repetitive bands that appear soon after the fusion of skeletal myoblasts into multinucleate fibers. This organization has been found in cultures containing myotubes that develop in vitro from explants of newborn rat hindlimb cells and in myotubes derived from the L8E63 myogenic line. Bands were also prominent in skinned fibers prepared from adult rat soleus muscle and in cardiac myocytes grown in vitro from 4-day heart ventricles. Immunofluorescence banding was localized in the sarcomere as a doublet, with one element on either side of the Z line. Evidence that supports the conclusion that the reaction with X22 antibody is specific and indicative of the localization of clathrin in the sarcomere includes: (1) Identical titration of X22 antibody reactivity with the determinant in coated vesicles and in the sarcomere. (2) Conditions (eg., pH and Tris) that disrupt clathrin baskets or prevent its assembly likewise disrupt the localization of X22 in bands. (3) Chloroquine inhibits both the normal trafficking of clathrin in the cell and X22 banding in the sarcomere. (4) Immunoblot analysis of myotube lysates reveals a single band with an electrophoretic mobility identical to the 180,000-Da clathrin heavy chain. (5) The assembly of clathrin into sarcomeric bands occurs early in the development of the myofibrillar apparatus. Quantitation of the appearance of X22 banding in primary cultures of myotubes indicates that it precedes that of other myofibrillar proteins and that assembly takes place in the following order: X22, titin, myosin heavy chain, actin, and desmin. The assembly of myosin, titin, and actin into sarcomeric bands, as well as X22, is inhibited by chloroquine. Upon prolonged exposure to chloroquine previously assembled proteins are drastically reduced or no longer evident in the sarcomere. On the basis of these results and considering the role of clathrin in intracellular transport and its capacity to interact with actin and alpha-actinin, we suggest that clathrin may have diverse roles in the assembly, integrity, and functioning of the sarcomere and its integration with the sarcolemma. The early organization of X22 into bands further suggests that clathrin may also function early in the assembly of the contractile system.     

Specific binding of sulphated polymers to ram sperm proacrosin

Sulphated polysaccharides and zona pellucida glycoproteins have been shown to bind non-enzymatically to proacrosin, the protein found within the acrosomal vesicle of mammalian spermatozoa. The mechanism of this interaction has been investigated using ¹²⁵I-fucoidan to probe purified ram sperm proacrosin. Results show that (a) binding of' ¹²⁵I-fucoidan to proacrosin is inhibited only by sulphated polymers and (b) recognition is mediated by poly(sulphate) groups and is largely independent of the composition of the polymer chain. It is suggested that a similar mechanism is responsible for the interaction between proacrosin and zona pellueida glycoproteins during the early stages of fertilization in mammals and this process mediates firm binding of spermatozoa to the egg.     

Proton-induced grana formation in chloroplasts. Distribution of chlorophyll-protein complexes and Photosystem II photochemistry

Lowering the pH of the incubation medium to pH 5.4 leads to grana formation morphologically similar to that induced by metal cations. The same phenomenon is observed in EDTA-washed chloroplasts, indicating that it is not due in part to electrostatic ‘masking’ by residual cations associated with the membranes. Digitonin fractionation studies have indicated that the distribution of the major chlorophyll-protein complexes between granal and stromal membrane regions is similar at pH 5.4 in the absence of Mg²⁺, and at pH 7.4 in the presence of Mg²⁺. Chlorophyll fluorescence induction studies have indicated that the primary photochemistry of Photosystem II (PS II) is stimulated by lowering the pH to 5.4, just as it is upon metal cation addition at higher pH values. The failure to observe such an increase at pH 5.4 by measuring electron transport to ferricyanide is attributed to a combination of an inhibition by this pH of electron transport at a site after Q reduction and an increase in the number of PS II centres detached from the plastoquinone pool. We conclude that the stacked configuration of chloroplast membranes leads to increased PS II primary photochemistry, which is most simply explained in terms of a redistribution of excitation energy towards PS II.     

The “High Irradiance Responses” of plant photomorphogenesis

Plants growing in the natural environment are exposed daily to prolonged periods of high intensity irradiation. Many plant photomorphogenic responses are fully expressed only under prolonged exposures to high irradiances of light. The intensive study of these responses, the “High Irradiance Responses” (HIR) of plant photomorphogenesis, which started about 20 years ago, has been essentially directed—so far—toward the identification of the HIR photoreceptor, or photoreceptors, a problem that has not been satisfactorily and definitively solved, as yet. There is a great deal of evidence in support of the hypothesis that phytochrome, the pigment mediating the redfar red reversible plant photo-responses to low fluences of light, is involved in the photocontrol of the HIR. It seems likely that phytochrome may be the only photomorphogenic receptor responsible for the photocontrol of HIR responses brought about by irradiation at wavelengths longer than 600 nm. Phytochrome is probably also involved in the photocontrol of the HIR effects brought about by irradiation in the 350 to 500 nm region of the spectrum, but it cannot be excluded that other photochemical systems may also be involved. From a theoretical point of view, it does not seem unreasonable that the final expression of an HIR response may involve an interaction between phytochrome and other photochemical systems, with phytochrome probably playing the primary role and being responsible for the control of the activity of the other systems. Numerous “phytochrome only” interpretations (models) of the HIR have been proposed. Some of them have been developed to a fairly high degree of elaboration and have allowed the prediction of at least some of the features of the HIR. These “models,” although not rigorously and completely tested yet, seem to provide a reasonable interpretation for the HIR effects displayed under prolonged far red irradiation and for those HIR responses for which far red is the most effective spectral region. However, they do not provide a satisfactory explanation for the HIR responses for which blue is the most or the only effective spectral region, nor for the high effectiveness of white light. But, in spite of these problems, the “phytochrome only” interpretations of the HIR can be considered more satisfactory than those based on an interaction between phytochrome and other photochemical systems, especially in relation to the fact that the identity of these other photochemical systems has not been defined yet.     

The proteins of the content of the secretory granules of the rat parotid gland

The proteins of the secretory granules of the rat parotid gland were characterized by sodium dodecylsulfate gel electrophoresis, by chromatography of [³H]prolinelabeled proteins on DEAE-cellulose and by amino acid analysis.Sodium dodecylsulfate gel electrophoresis of the secretory granule content showed five principal proteins and a limited number of minor components. Only two of the principal bands could be identified as known secretory enzymes of the parotid gland. One was identified as the α-amylase and one as deoxyribonuclease. Peroxidase and ribonuclease form minor portions of the secretory proteins.The other three major proteins constitute, together, about 60% by weight, of the secretory granule content proteins. Of these, one which represents more than 30% of the total granule protein was found to contain uniquely high amounts of leucine residues (21 mole%). Another one of these principal proteins was relatively rich in cysteine residues (7 mole%).The fifth principal protein was found to contain high amounts of proline (28 mole%) glutamic acid (17 mole%) and glycine (18 mole%) residues. Its amino acid composition was very similar to that of the proline-rich proteins that were previously shown to be present in the membrane isolated from these granules. This protein, however, differed from the “membranous” proline-rich proteins by several criteria.Two minor glycoproteins of the secretory granule content were also found to be rich in proline residues (37 mole%). As with the other proline-rich proteins of the granule, they contained no sulphur-containing amino acids, stained faintly pink with Coomassie Blue and were underestimated by the Lowry method. They differ however, from all the other proline-rich proteins of the granule by having a significantly higher content of threonine, less glycine (9 mole%) and much less glutamic acid (3 mole%).Of the principal proteins, only the deoxyribonuclease and the half-cystine-rich proteins were positively stained by periodic acid Schiff staining.The possible functions of the leucine-rich, the half cystine-rich and the various proline-rich proteins are discussed.     

Basic protein changes during the final stages of sperm maturation in the house cricket

Polyacrylamide gel electrophoresis has been used to analyse basic protein changes during the final stages of spermiogenesis in the house cricket. Mature sperm were obtained from the spermathecae of inseminated females. Their basic protein is electrophoretically heterogeneous, with two major and two minor components, all of unusually high mobilities, as expected ofprotamine. No histones are present. Testis also contains basic protein components of high mobilities, although in small amount relative to the histones present. Testis preparations were centrifuged on a density gradient of colloidal silica to separate nuclei of different stages of spermiogenesis from each other, and it was found that very late spermatids contain relatively large amounts of protamine. At least seven different protamine-like components, each with a different mobility, occur during the final maturation stages. The particular components present, and their abundancies, vary during development. The complement first found in spermatids is different from that of a later spermatid; still another complement is found in sperm from the seminal vesicle; and still another in mature sperm. Components which are abundant in spermatids are progressively eliminated, while components which are barely detectable in them gradually increase in abundance to become the major components of the basic protein complement at maturity.     

Uptake and fate of IAA in apple callus tissues: metabolism of IAA-2-14C

Young and old apple callus tissues were incubated in light ordarkness with IAA-2-¹⁴C. A large portion of the IAA disappearedfrom the medium with both young and old calluses. Whereas withold calluses the loss was mainly due to IAA destruction, youngcalluses accumulated IAA to a level which exceeded the externalconcentration and, in addition, seemed to protect it from breakdown.After 24 hr the level of IAA-2-¹⁴C in the medium dropped to50% with old calluses both in the dark and light, and with youngcalluses to 20% in the light and 50% in the dark. Chromatographyand scanning of the media and calluses showed that IAA was convertedinto two compounds (comp. A and comp. B). The amounts and proportionsof these metabolites in the medium and tissue were dependenton the different treatments and callus age. The breakdown ofIAA by old tissue gave rise to a higher level of comp. B bothin the tissue and medium, particularly after 6 hr of incubation.In the medium of young tissues the level of comp. A was higherthan comp. B while equal amounts of the two compounds were detectedin the tissue, itself. The origin of the IAA products in thetissue was probably endogenous and not via absorption from themedium. The IAA metabolism of apple callus tissues seems toproceed via the oxindole pathway and it is proposed that compoundsA and B are 3-hydroxymethyloxindole and 3-methylene oxindole,respectively. ¹ Contribution from the Agricultural Research Origanization,The Volcani Center, Bet Dagan, Israel. 1973 Series No. 275-E. (Received May 30, 1974; )