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1 Pasteur Institute-Cenci Bolognetti Foundation, Department of Cell and Developmental Biology, University of Rome ‘La Sapienza’, Rome, Italy
2 Department of Experimental Medicine and Pathology, University of Rome ’La Sapienza’, Rome, Italy
ABSTRACT
The P-type Ca2+-ATPases are the transporters responsible for calcium homeostasis in the cell compartments of eukaryotes. The KlPMR1 gene of Kluyveromyces lactis encodes a P-type Ca2+-ATPase, which is functionally and structurally homologous to Pmr1p of Saccharomyces cerevisiae, the calcium pump localized in the Golgi membranes. In this work, a novel involvement of KlPmr1p in cell-wall morphogenesis of K. lactis is reported. Klpmr1 
cells exhibited the loss of outer-chain extension in the glycosylation of secreted proteins. The absence of KlPmr1p resulted in the accumulation of round, large cells with an abnormally thick cell wall, as revealed by transmission electron microscopy. The deletant strain also showed a delocalized deposition of chitin in the lateral cell wall accompanied by an unbalanced ratio of insoluble to soluble glucans. These morphological defects were accompanied by the presence of irregularly shaped nuclei and by a DNA content greater than 2n. Addition of 10 mM Ca2+ to the medium of the Klpmr1 strain reversed the chitin-deposition impairment, recovered the alteration to the glucan ratio and restored a normal thickness of the cell wall. The mutant cells resumed wild-type size, shape and nuclear morphology but the DNA content indicated the persistence of defects in the co-ordination between DNA replication and cell division. The glycosylation defects were completely unaffected by the calcium supplement. These results indicate that calcium homeostasis controlled by KlPmr1p plays an important role in the cell-wall morphogenesis of K. lactis.
Author for correspondence: Claudio Palleschi. Tel: +39 0649912321. Fax: +39 0649912351. e-mail: palleschic@axcasp.caspur.it
Present address: Department of Developmental and Cell Biology, University of Rome 1, Piazzale Aldo Moro, 5-00185 Rome, Italy.
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