Rosa Lacalle - Academia.edu (original) (raw)

Papers by Rosa Lacalle

Molecular and Cellular Biology, Apr 1, 1999

Iubmb Life, 2000

Throughout evolution, both prokaryotic and eukaryotic cells have developed a variety of biochemic... more Throughout evolution, both prokaryotic and eukaryotic cells have developed a variety of biochemical mechanisms to de ne the direction and proximity of extracellular stimuli. This process is essential for the cell to reply properly to the environmental cues that determine cell migration, proliferation, and differentiation. Chemotaxis is the cellular response to chemical attractants that direct cell migration, a process that plays a central role in many physiological situations, such as host immune responses, angiogenesis, wound healing, embryogenesis, and neuronal patterning, among others. In addition, cell migration takes part in pathological states, including in ammation and tumor metastasis. Indeed, tumor progression to invasion and metastasis depends on the active motility of the invading cancer cells and the endothelial cell bed during tumor neovascularization. Cell migration switches "off" and "on," based on quantitative differences in molecular components such as adhesion receptors, cytoskeletal linking proteins, and extracellular matrix ligands, and by regulating the af nity of membranebound chemoattractant receptors. A clear understanding of how cells sense chemoattractants is, therefore, of pivotal importance in the biology of the normal cell as well as in prevention of malignant cell invasion. Here we offer a perspective on cell migration that emphasizes the relationship between cell polarization and cell movement and the importance of the equilibrium between the signals that drive each process for the control of tumor cell invasion. IUBMB Life, 49: 89 -96, 2000

Trends in Immunology, Jan 6, 2003

Seminars in immunology, 2001

Cell chemotaxis requires the acquisition and maintenance of both spatial and functional asymmetry... more Cell chemotaxis requires the acquisition and maintenance of both spatial and functional asymmetry between initially equivalent cell parts. In leukocytes one becomes the leading edge and the other, the rear edge or uropod. The acquisition of this cell polarity is controlled by an array of chemoattractants, including those of the chemokine family. We propose that chemokine receptor activation in highly organized lipid raft domains is a major determinant for the correct localization of the signaling pathways leading to the cell asymmetries required for migration. The lateral organization imposed by membrane raft microdomains is discussed in the context of other chemokine receptor activities, such as its role as a human immunodeficiency virus (HIV) coreceptor.

Molecular and cellular biology, 1999

The coordinated interplay of substrate adhesion and deadhesion is necessary for cell motility. Us... more The coordinated interplay of substrate adhesion and deadhesion is necessary for cell motility. Using MCF-7 cells, we found that insulin-like growth factor I (IGF-I) induces the adhesion of MCF-7 to vitronectin and collagen in a dose- and time-dependent manner, suggesting that IGF-I triggers the activation of different integrins. On the other hand, IGF-I promotes the association of insulin receptor substrate 1 with the focal adhesion kinase (FAK), paxillin, and the tyrosine phosphatase SHP-2, resulting in FAK and paxillin dephosphorylation. Abrogation of SHP-2 catalytic activity with a dominant-negative mutant (SHP2-C>S) abolishes IGF-I-induced FAK dephosphorylation, and cells expressing SHP2-C>S show reduced IGF-I-stimulated chemotaxis compared with either mock- or SHP-2 wild-type-transfected cells. This impairment of cell migration is recovered by reintroduction of a catalytically active SHP-2. Interestingly, SHP-2-C>S cells show a larger number of focal adhesion contacts ...

Cancer Metastasis - Biology and Treatment, 2004

Page 1. 1. OVERVIEW Progression from a benign, non-invasive in situ carcinoma to a malignant, inv... more Page 1. 1. OVERVIEW Progression from a benign, non-invasive in situ carcinoma to a malignant, invasive breast carcinoma is a complex multistage process, in which uncontrolled cell proliferation is a hallmark of the disease. ...

Trends in Immunology, 2003

The Journal of Cell Biology, 2002

ell signaling does not occur randomly over the cell surface, but is integrated within cholesterol... more ell signaling does not occur randomly over the cell surface, but is integrated within cholesterol-enriched membrane domains, termed rafts. By targeting SHP-2 to raft domains or to a non-raft plasma membrane fraction, we studied the functional role of rafts in signaling. Serumdepleted, nonattached cells expressing the raft SHP-2 form, but not non-raft SHP-2, display signaling events resembling those observed after fibronectin attachment, such as ␤ 1 integrin clustering, 397 Y-FAK phosphorylation, and ERK activation, and also increases Rho-GTP levels. Expression of C the dominant negative N19Rho abrogates raft-SHP-2-induced signaling, suggesting that Rho activation is a downstream event in SHP-2 signaling. Expression of a catalytic inactive SHP-2 mutant abrogates the adhesion-induced feedback inhibition of Rho activity, suggesting that SHP-2 contributes to adhesion-induced suppression of Rho activity. Because raft recruitment of SHP-2 occurs physiologically after cell attachment, these results provide a mechanism by which SHP-2 may influence cell adhesion and migration by spatially regulating Rho activity.

The Journal of Cell Biology, 2004

patially restricted activation of signaling molecules governs critical aspects of cell migration;... more patially restricted activation of signaling molecules governs critical aspects of cell migration; the mechanism by which this is achieved nonetheless remains unknown. Using time-lapse confocal microscopy, we analyzed dynamic redistribution of lipid rafts in chemoattractant-stimulated leukocytes expressing glycosyl phosphatidylinositolanchored green fluorescent protein (GFP-GPI). Chemoattractants induced persistent GFP-GPI redistribution to the leading edge raft (L raft) and uropod rafts of Jurkat, HL60, and dimethyl sulfoxide-differentiated HL60 cells in a pertussis toxin-sensitive, actin-dependent manner. A trans-S membrane, nonraft GFP protein was distributed homogeneously in moving cells. A GFP-CCR5 chimera, which partitions in L rafts, accumulated at the leading edge, and CCR5 redistribution coincided with recruitment and activation of phosphatidylinositol-3 kinase ␥ in L rafts in polarized, moving cells. Membrane cholesterol depletion impeded raft redistribution and asymmetric recruitment of PI3K to the cell side facing the chemoattractant source. This is the first direct evidence that lipid rafts order spatial signaling in moving mammalian cells, by concentrating the gradient sensing machinery at the leading edge.

Seminars in Immunology, 2005

Many immune cells can detect the direction and intensity of an extracellular chemical gradient, a... more Many immune cells can detect the direction and intensity of an extracellular chemical gradient, and migrate toward the source of stimulus. This process, called chemotaxis, is essential for immune system function and homeostasis, and its deregulation is associated with serious diseases. Chemotaxis is initiated by chemoattractant binding to heterotrimeric G protein-coupled receptors, which translate the gradients into accurate directional migration. A necessary step in this process is cell polarization, the acquisition of functional and spatial asymmetry. The use of new imaging technologies enables analysis of spatial and temporal changes in the activity of proteins and membrane domains involved in polarization and chemotaxis. We discuss the sometimes contradictory evidence available and the emerging molecular model for immune cell polarity and chemotaxis.

Journal of Cell Science, 2004

IUBMB Life (International Union of Biochemistry and Molecular Biology: Life), 2000

EMBO reports, 2001

Complex cell responses require the integration of signals delivered through different pathways. W... more Complex cell responses require the integration of signals delivered through different pathways. We show that insulin-like growth factor (IGF)-I induces specific transactivation of the G i -coupled chemokine receptor CCR5, triggering its tyrosine phosphorylation and Gα i recruitment. This transactivation occurs via a mechanism involving transcriptional upregulation and secretion of RANTES, the natural CCR5 ligand. CCR5 transactivation is an essential downstream signal in IGF-I-induced cell chemotaxis, as abrogation of CCR5 function with a transdominant-negative KDELccr5∆32 mutant abolishes IGF-Iinduced migration. The relevance of this transactivation pathway was shown in vivo, as KDELccr5∆32 overexpression prevents invasion by highly metastatic tumor cells; conversely, RANTES overexpression confers built-in invasive capacity on a non-invasive tumor cell line. Our results suggest that this extracellular growth factor-chemokine network represents a general mechanism connecting tumorigenesis and inflammation.

The Embo Journal, Nov 15, 1999

The acquisition of spatial and functional asymmetry between the rear and the front of the cell is... more The acquisition of spatial and functional asymmetry between the rear and the front of the cell is a necessary step for cell chemotaxis. Insulin-like growth factor-I (IGF-I) stimulation of the human adenocarcinoma MCF-7 induces a polarized phenotype characterized by asymmetrical CCR5 chemokine receptor redistribution to the leading cell edge. CCR5 associates with membrane raft microdomains, and its polarization parallels redistribution of raft molecules, including the raft-associated ganglioside GM1, glycosylphosphatidylinositol-anchored green fluorescent protein and ephrinB1, to the leading edge. The non-raft proteins transferrin receptor and a mutant ephrinB1 are distributed homogeneously in migrating MCF-7 cells, supporting the raft localization requirement for polarization. IGF-I stimulation of cholesterol-depleted cells induces projection of multiple pseudopodia over the entire cell periphery, indicating that raft disruption specifically affects the acquisition of cell polarity, but not IGF-I-induced protrusion activity. Cholesterol depletion inhibits MCF-7 chemotaxis, which is restored by replenishing cholesterol. Our results indicate that initial segregation between raft and non-raft membrane proteins mediates the necessary redistribution of specialized molecules for cell migration.

Endocrinology, May 1, 1999

MCF-7 cells migrate through vitronectin-coated filters in response to insulin-like growth factor ... more MCF-7 cells migrate through vitronectin-coated filters in response to insulin-like growth factor I (IGF-I); migration is inhibited by the matrix metalloproteinase (MMP) inhibitor BB-94, but not by the serine proteinase inhibitor aprotinin. MMP-9 was identified in the conditioned medium of MCF-7 cells; in addition, fluorescence-activated cell sorting analysis revealed its presence on the cell surface, where MMP-9 activity was also found using a specific fluorogenic peptide. Furthermore, the messenger RNA encoding MMP-9 was detected in MCF-7 cells by PCR. The IGF-I concentration leading to maximal MCF-7 invasion produces an increase in cell surface pro-teolytic activity after short incubation periods. At 18 h, however, preincubation of MCF-7 cells with IGF-I produces at 18 h a dosedependent decrease in cell-associated MMP-9 activity and an increase in soluble MMP-9. MCF-7 invasion is dependent on the ␣ v ␤ 5 integrin, a vitronectin receptor. The levels of ␣ v -and ␤ 5 -subunits expressed in MCF-7 cells depend on the IGF-I concentration, which triggers an increase in both of these subunits. Based on these results, we suggest that IGF-I-induced MCF-7 cell migration is mediated by the MMP-9 activity on the cell surface and by ␣ v ␤ 5 integrin. (Endocrinology 140: [1657][1658][1659][1660][1661][1662][1663][1664] 1999)

EMBO Reports

We show that these membrane changes are necessary for infection, as initial gp120-CD4 engagement ... more We show that these membrane changes are necessary for infection, as initial gp120-CD4 engagement leads to redistribution and clustering of membrane microdomains, enabling subsequent interaction of this complex with HIV-1 co-receptors. Disruption of cell membrane rafts by cholesterol depletion before viral exposure inhibits entry by both X4 and R5 strains of HIV-1, although viral replication in infected cells is unaffected by this treatment. This inhibitory effect is fully reversed by cholesterol replenishment of the cell membrane. These results indicate a general mechanism for HIV-1 envelope glycoprotein-mediated fusion by reorganization of membrane microdomains in the target cell, and offer new strategies for preventing HIV-1 infection.

Clinical & experimental metastasis, 2002

Tumor cells acquire the ability to enter blood vessels surrounding the primary tumor, endowing th... more Tumor cells acquire the ability to enter blood vessels surrounding the primary tumor, endowing them with the capacity to disseminate and become established in distant sites, originating a metastasis. Determination of the intravasation ability of tumor cells is thus important, as it can be correlated with their potential malignancy. To analyze the intravasation phenotype of human tumor cells in vivo, we performed chick embryo chorioallantoic membrane (CAM) assays. Cells were inoculated on the CAM of 9-day-old chick embryos and the membrane at the opposite side of the egg was recovered after 48 h incubation. To measure intravasation ability, we calculated the amount of human DNA in each CAM sample by real-time PCR of Alu sequences and SYBR Green 1 fluorescence detection. This analysis showed a detection limit of 1 human cell per 10(5) total cells, and we were able to distinguish between tumor cells of distinct invasive capacity. This assay has several advantages over current methods t...

Journal of bacteriology, 1993

The biologically inactive compound N-acetylpuromycin is the last intermediate of the puromycin an... more The biologically inactive compound N-acetylpuromycin is the last intermediate of the puromycin antibiotic biosynthetic pathway in Streptomyces alboniger. Culture filtrates from either this organism or Streptomyces lividans transformants harboring the puromycin biosynthetic gene cluster cloned in low-copy-number cosmids contained an enzymic activity which hydrolyzes N-acetylpuromycin to produce the active antibiotic. A gene encoding the deacetylase enzyme was located at one end of this cluster, subcloned in a 2.5-kb DNA fragment, and expressed from a high-copy-number plasmid in S. lividans.