RESUMEN
The structural complexity of glycosylation restrains the functional characterization of glycans. We present a versatile carbohydrate ligation technique based on the reaction of cyclic carbamates with primary amines. Cyclic-carbamate-derivatized carbohydrates can be added to primary amine-containing molecules in aqueous solution to yield glycoconjugates. This method enabled the presentation of carbohydrate epitopes on live animal cells, as shown by the acquisition of E-selectin binding sites on mouse MC-38 cells decorated with 3-fucosyllactose or 3-fucosyl-3-sialyllactose. Ligation of 3- and 6-sialyllactose to Escherichia coli demonstrated the importance of sialic acid linkages in regulating complement factor H binding. Proteins were modified with oligosaccharides to study their role in stimulating cytokine secretion by dendritic cells, thus pointing to interactions between glycoproteins and phosphoinositide 3-kinase signaling in controlling interleukin-12, tumor necrosis factor alpha and interleukin-1ß release. Overall, cyclic-carbamate-mediated ligation is useful to study the biology of carbohydrate epitopes on proteins and on cell membranes.
Asunto(s)
Carbamatos/química , Pared Celular/química , Escherichia coli/metabolismo , Oligosacáridos/química , Animales , Bovinos , Línea Celular , Pared Celular/inmunología , Pared Celular/metabolismo , Células Dendríticas/citología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Selectina E/química , Selectina E/metabolismo , Epítopos/química , Epítopos/inmunología , Glicosilación , Humanos , Interleucina-12/metabolismo , Interleucina-1beta/metabolismo , Ratones , Oligosacáridos/síntesis química , Oligosacáridos/inmunología , Fosfatidilinositol 3-Quinasas/metabolismo , Albúmina Sérica Bovina/química , Albúmina Sérica Bovina/metabolismo , Transducción de Señal , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The life cycle of flowering plants is marked by several post-embryonic developmental transitions during which novel cell fates are established. Notably, the reproductive lineages are first formed during flower development. The differentiation of spore mother cells, which are destined for meiosis, marks the somatic-to-reproductive fate transition. Meiosis entails the formation of the haploid multicellular gametophytes, from which the gametes are derived, and during which epigenetic reprogramming takes place. Here we show that in the Arabidopsis female megaspore mother cell (MMC), cell fate transition is accompanied by large-scale chromatin reprogramming that is likely to establish an epigenetic and transcriptional status distinct from that of the surrounding somatic niche. Reprogramming is characterized by chromatin decondensation, reduction in heterochromatin, depletion of linker histones, changes in core histone variants and in histone modification landscapes. From the analysis of mutants in which the gametophyte fate is either expressed ectopically or compromised, we infer that chromatin reprogramming in the MMC is likely to contribute to establishing postmeiotic competence to the development of the pluripotent gametophyte. Thus, as in primordial germ cells of animals, the somatic-to-reproductive cell fate transition in plants entails large-scale epigenetic reprogramming.