Logo Università degli Studi di Milano



 
 

Laboratory of Membrane physiology  

Lab. Sacchi

 

Main Investigator:  Prof. Vellea Franca Sacchi  Franca.Sacchi@unimi.it   

Team
Michela Castagna Assistant Professor        Michela.Castagna@unimi.it
Matteo Giovanola PhD student                  Matteo.Giovanola@unimi.it
Elena Signoretto PhD student                   Elena.Signoretto@unimi.it

 

Research topics

Introduction:

The laboratory has an established experience in structural and functional characterization of neurotransmitter and amino acid transporters. The attention is focused on members of the NSS/SLC6 family that play a key role in the termination of synaptic transmission and in intestinal and renal solute absorption and reabsorption. We are also interested in the study of iron transporters that have an essential function in the complex pathway that regulates uptake, storage, utilization and  secretion of this metabolically relevant ion.

Research topic outline:

  1. Structure-function analysis of NSS/SLC6 neurotransmitter and amino acid transporters with the aim of the identification of structural determinants of interaction with substrates, inhibitors and regulatory proteins and the understanding of mechanisms controlling transport activity and cellular localization of the proteins
  2. Molecular genetic analysis of eukaryotic iron homeostasis
  3. Biophysical characterization of  water transport in solute cotransporters
  4. Atomic force microscopy (AFM) visualization of membrane proteins

 

Techniques

  • Molecular biology techniques for cloning, expression and site-directed mutagenesis  of membrane proteins and solute cotransporters
  • Functional analysis of transporters expressed in heterologous systems such as Xenopus laevis oocytes and stable cell lines or purified in brush border membrane vesicles
  • Radioactive solute flux measurements, 3H-thimidine and 35S incorporation, SDS page, western-blot
  • Preparation of Xenopus oocytes plasma membrane specimens for visualization by atomic force microscopy (AFM)

 

Selected publications

  1. M. Giovanola, F. D’Antoni, M. Santacroce, S.A. Mari, F. Cherubino, E. Bossi, V.F. Sacchi and M. Castagna (2012) Role of a conserved glycine triplet in the NSS amino acid transporter KAAT1. BBA-Biomembranes. 1818: 1737-1744. I.F (2011): 3.990.
  2. S. Bertram, F. Cherubino, E. Bossi , M. Castagna, A. Peres. (2011) GABA reverse transport by the neuronal cotransporter GAT1: influence of internal chloride depletion. Am. J. Physiol Cell Physiol. 301: C1064-C1073. I.F.: 3.536.
  3. M. Santacroce, M. Castagna and V.F. Sacchi (2010) Passive water permeability of some wild type and mutagenized amino acid cotransporters of the SLC6/NSS family expressed in Xenopus laevis oocytes. Comp. Biochem. Physiol. A Mol. & Integr. Physiol 156: 509-517.IF.2.134.
  4. F. Orsini, M. Santacroce, P. Arosio, M. Castagna, C. Lenardi, G. Poletti, V.F. Sacchi. (2009) Intermittent contact mode AFM investigation of native plasma membrane of Xenopus laevis oocyte. European Biophysics Journal.38: 903-910. I.F. 2.437.
  5. M. Castagna, A. Soragna, S.A. Mari, M. Santacroce, S. Bettè, P.G. Mandela, G. Rudnick, A. Peres, V.F. Sacchi. (2007). Interaction between lysine 102 and aspartate 338 in the insect amino acid cotransporter KAAT1. Am. J. Physiol. Cell. Physiol. 293: C1286-C1295. I.F. 4.230.
  6. Miszner, A. Peres, M. Castagna, S. Bettè, S. Giovannardi, F. Cherubino, E. Bossi.(2007) Structural and functional basis of amino acid specificity in the invertebrate cotransporter KAAT1.  J. of Physiol. 581: 899-931.  I.F. 4.58
  7. S.A. Mari, A. Soragna, M. Castagna, M. Santacroce, E. Bossi, A. Peres, V.F. Sacchi. (2006) Role of the conserved glutamine 291 in the rat g-aminobutyric acid transporter rGAT-1. Cell. Mol. Life Sci. 63, 100-111. I.F. 4.655.
  8. S.A. Mari, A. Soragna, M. Castagna, E. Bossi, A. Peres, V.F. Sacchi  (2004) Aspartate 338 contributes to the cationic specificity and to the driver-amino acid coupling in the insect cotransporter KAAT1. Cell. Mol. Life Sci. 61: 243-256. IF 4.812
  9. E. Bossi, E. Centinaio,  M. Castagna, S. Giovannardi, S. Vincenti, V.F. Sacchi, A. Peres (1999) Ion binding and permeation trough the lepidopteran amino acid transporter KAAT1 expressed in Xenopus oocytes. J. of Physiology 515.3, 729-742. I.F. 4.650.
  10. M. Castagna, C. Shayakul, D. Trotti, V.F. Sacchi, W.R. Harvey, M.A. Hediger (1998) Cloning and characterization of a potassium-coupled amino acid transporter. Proc. Natl. Acad. Sci. 95, 5395-5400. I.F 10.700.
Back to top