G M1 gangliosidosis is an inherited, fatal neurodegenerative disease caused by deficiency of lysosomal β-D-galactosidase (EC 3.2.1.23) and consequent storage of undegraded G M1 ganglioside. To characterize the genetic mutation responsible for feline G M1 gangliosidosis, the normal sequence of feline β-galactosidase cDNA first was defined. The feline β-galactosidase open reading frame is 2010 base pairs, producing a protein of 669 amino acids. The putative signal sequence consists of amino acids 1-24 of the β-galactosidase precursor protein, which contains seven potential N-linked glycosylation sites, as in the human protein. Overall sequence homology between feline and human β-galactosidase is 74% for the open reading frame and 82% for the amino acid sequence. After normal β-galactosidase was sequenced, the mutation responsible for feline G M1 gangliosidosis was defined as a G to C substitution at position 1448 of the open reading frame, resulting in an amino acid substitution at arginine 483, known to cause G M1 gangliosidosis in humans. Feline β-galactosidase messenger RNA levels were normal in cerebral cortex, as determined by quantitative RT-PCR assays. Although enzymatic activity is severely reduced by the mutation, a full-length feline β-galactosidase cDNA restored activity in transfected G M1 fibroblasts to 18-times normal. β-Galactosidase protein levels in G M1 tissues were normal on Western blots, but immunofluorescence analysis demonstrated that the majority of mutant β-galactosidase protein did not reach the lysosome. Additionally, G M1 cat fibroblasts demonstrated increased expression of glucose-related protein 78/BiP and protein disulfide isomerase, suggesting that the unfolded protein response plays a role in pathogenesis of feline G M1 gangliosidosis. CIHR Author Manuscript CIHR Author Manuscript CIHR Author ManuscriptThe lysosomal enzyme β-D-galactosidase (βgal, EC 3.2.1.23) cleaves terminal galactose residues from a variety of molecules, including gangliosides G A1 and G M1 . Deficiency of βgal is known to cause two lysosomal storage diseases: G M1 gangliosidosis (neuronopathic) and Morquio B Disease (mucopolysaccharidosis IVB, non-neuronopathic [3,4]. Although the in vivo biochemical effect of the Arg482His mutation often is difficult to discern because it occurs most frequently in compound heterozygotes, patients homozygous for the G1445A substitution present with the infantile (most severe) form of G M1 gangliosidosis [3,7,8]. A similar mutation, Arg482Cys, also produced no residual βgal activity after expression in G M1 gangliosidosis fibroblasts [4].Feline G M1 gangliosidosis, first described in a Siamese cat in 1971 [9], models the juvenile form of the human disease. Onset of clinical neurological disease in affected cats occurs at approximately 3.5 months of age with a fine head or limb tremor. G M1 mutant cats have progressive dysmetria and ambulatory difficulties, with blindness and epileptiform seizures in the terminal disease stage at 9-10 months of age. In the current st...
In our phylogenetic analysis of the subfamily Pitcairnioideae, three monophyletic groups of genera became evident. These are formally treated as tribes, namely, Brocchinieae (Brocchinia); Pitcairnieae (Ayensua, and Steyerbromelia); Puyeae (Abromeitiella, Brewcaria, Deuterocohnia, Dyckia, Encholirium, Hechtia, and Puya). Pepinia, most recently considered a subgenus of Pitcairnia, is recognized as a genus in Pitcairnieae.
Scanning electron and light microscopy observations of wet-preserved flowers of Bromeliaceae subfamily Pitcairnioideae yield new information on the stigma, petal scales, and septal nectaries. Variations of the stigma types are evident among several genera. The gross structural features of the stigma do not indicate definite pollination trends, but the shape of the lobes and papillae indicate a few specific modes. In pitcairnioid genera, petal scales, when functional, may aid in pollination by accumulating the nectar secreted from the ovary, thus facilitating its availability to the pollinator. Nectaries associated with the gynoecia usually display tripartite channels in the ovary septa. Some developmental changes of the channel structure and position of the ovary indicate three probable modes of nectar release from the gynoecia of the pitcairnioids: (1) through lateral grooves or openings, (2) partly through the apical orifices and partly through the dissolved areas of the spetal channels, and (3) exclusively through the apical orifices. Analysis of a wide range of floral features indicates that ornithophily, chiropterophily, and entomophily exist in different Pitcairnioideae lineages. Gynoecia of the monocotyledons often contain nectaries in the ovary septa (septal nectaries) and are known to be of taxonomic value in several families (DAUMANN 1970; FAHN 1982; THORNE 1983; DAHLGREN et al. 1985). BUDNOWSKI S (1922) examination represents the only major study of Bromeliaceae septal nectaries. Their morphological variation and the significance in pollination were not analyzed until the present work. The goal of this study is to examine the diversity of stigma morphology, petal scales, and septal nectaries of the subfamily Pitcairnioideae from wetpreserved material. Material and methods The 41 species of nine genera of Pitcairnioideae were examined by light microscopy; 30 species were also examined with scanning electron microscopy (SEM) (table 1). All material was field-collected by G. S. VARADARAJAN and preserved in FAA (formaldehyde:acetic acid:ethanol, 1: 1: 18). Samples of stigmas and ovaries were selected from preanthesis to post-anthesis, and the internal structure of ovaries was examined from freehand cross sections. Petal scales were examined from nearly mature (anthesis) flowers. Floral parts selected for SEM were transferred from FAA to 0.1 M cacodylate buffer, postfixed in OSO4, dehydrated through an ethanolic series (30%, 50%, 70%, 95%, 100%), dried in a Bomar critical-point dryer, mounted on aluminum stubs, and gold coated with a Technics Hummer-Sputter coater. Flower samples were then examined in an ETEC Auto Scan U-1 scanning electron microscope at 20 kV.
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