[Fragility in a Public Health perspective: principles and tools for a "life course" approach prevention- oriented]. / La Fragilità in una prospettiva di Sanità Pubblica: principi e strumenti per un approccio "life course" orientato alla prevenzione.

The Covid-19 pandemic significantly increased the workload for the Italian Health Service. There is few information in the literature on the pediatric population and on the management of pediatric hospitals. The aim of this article is to describe the management of healthcare services during Covid-19 emergency in Regina Margherita Children's Hospital. The Regina Margherita Children's Hospital is specialized in the prevention, diagnosis and treatment of pediatric diseases. About 1000 health worker work in this Hospital and 278 hospitalization places are available.

Small molecules targeted to the microtubule-Hec1 interaction inhibit cancer cell growth through microtubule stabilization.

Highly expressed in cancer protein 1 (Hec1) is a subunit of the kinetochore (KT)-associated Ndc80 complex, which ensures proper segregation of sister chromatids at mitosis by mediating the interaction between KTs and microtubules (MTs). HEC1 mRNA and protein are highly expressed in many malignancies as part of a signature of chromosome instability. These properties render Hec1 a promising molecular target for developing therapeutic drugs that exert their anticancer activities by producing massive chromosome aneuploidy. A virtual screening study aimed at identifying small molecules able to bind at the Hec1-MT interaction domain identified one positive hit compound and two analogs of the hit with high cytotoxic, pro-apoptotic and anti-mitotic activities. The most cytotoxic analog (SM15) was shown to produce chromosome segregation defects in cancer cells by inhibiting the correction of erroneous KT-MT interactions. Live cell imaging of treated cells demonstrated that mitotic arrest and segregation abnormalities lead to cell death through mitotic catastrophe and that cell death occurred also from interphase. Importantly, SM15 was shown to be more effective in inducing apoptotic cell death in cancer cells as compared to normal ones and effectively reduced tumor growth in a mouse xenograft model. Mechanistically, cold-induced MT depolymerization experiments demonstrated a hyper-stabilization of both mitotic and interphase MTs. Molecular dynamics simulations corroborate this finding by showing that SM15 can bind the MT surface independently from Hec1 and acts as a stabilizer of both MTs and KT-MT interactions. Overall, our studies represent a clear proof of principle that MT-Hec1-interacting compounds may represent novel powerful anticancer agents.

Outcomes of the calcaneo-stop procedure for the treatment of juvenile flatfoot in young athletes.

PURPOSE: Flexible flatfoot (FFF) is a widespread condition in juvenile patients. If symptomatic, FFF can require surgical treatment. The calcaneo-stop procedure has shown excellent clinical and radiographic outcomes and low rates of complications. The aim of the present study was to assess the sport practice of young athletes affected by FFF having undergone the calcaneo-stop procedure. METHODS: Between 2008 and 2016, 68 sport practitioners were bilaterally treated by the calcaneo-stop procedure, for a total of 136 FFF cases. Clinical evaluation, including the American Orthopedic Foot and Ankle Score (AOFAS), the Yoo et al score and The Foot & Ankle Disability Index (FADI) and FADI Sport scores were assessed. Radiographic evaluation was based on measurement of talar declination, Costa-Bertani's angle and calcaneal pitch. RESULTS: Mean follow-up was 57.6 months (sd 16.8). The AOFAS score mean increased from 79.3 (sd 5.7) to 97.3 (sd 4.5) three years after surgery. The Yoo score improved from 3.1 (sd 1.0) preoperatively to 11.7 (sd 0.6) three years after surgery. The FADI Sport subscale mean improved from 74.1 (sd 10.4) preoperatively to 95.9 (sd 4.9) three years after surgery.Costa-Bertani's angle decreased from 156.1° (sd 4.2°) to 135.8° (sd 7.3°) at three years postoperatively; mean talar declination angle decreased from 44.2° (sd 6.3°) to 30.6° (sd 3.2°) at three years postoperatively and mean calcaneal pitch increased from 12.6° (sd 2.3°) to 16.3° (sd 1.3°) at three years postoperatively. CONCLUSION: Adolescent patients who underwent the calcaneo-stop procedure reported satisfactory outcomes in terms of clinical and radiological evaluations. Moreover, our results showed an improvement of sport activity levels, with patients recovering sports activity within three months of surgery and without limitation in the execution of preferred activities. LEVEL OF EVIDENCE: IV.

NORMAL NUTRITIONAL COMPONENTS AND EFFECTS ON BONE METABOLISM IN PREVENTION OF OSTEOPOROSIS.

Osteoporosis is the most common bone disease, affecting millions of people and causing a high risk of fractures and a loss of quality of life. It is characterized by low bone mass and microarchitectural deterioration of bone tissue, with a consequent increase in bone fragility and susceptibility to fracture. A primary method of prevention, in order to reduce the risk of fractures, is represented by an appropriate lifestyle and a correct diet. There are potentially numerous nutrients and dietary components that can influence bone health, and these range from macronutrients to micronutrients as well as bioactive food ingredients. The purpose of this review is to overview osteoporosis, including its definition, etiology, and incidence, and then provide some information on possible dietary strategies for optimizing bone health and preventing osteoporosis. A correct diet to prevent osteoporosis should contain adequate amounts of calcium, vitamins D and K, protein, and fatty acids. The effects of these elements are briefly discussed, reporting on their correlation with bone benefits.

Performance of the CLS Spotorno uncemented stem in the third decade after implantation.

In 2012 we reviewed a consecutive series of 92 uncemented THRs performed between 1986 and 1991 at our institution using the CLS Spotorno stem, in order to assess clinical outcome and radiographic data at a minimum of 21 years. The series comprised 92 patients with a mean age at surgery of 59.6 years (39 to 77) (M:F 43;49). At the time of this review, seven (7.6%) patients had died and two (2.2%) were lost to follow-up. The 23-year Kaplan-Meier survival rates were 91.5% (95% confidence intervals (CI) 85.4% to 97.6%; 55 hips at risk) and 80.3% (95% CI, 71.8% to 88.7%; 48 hips at risk) respectively, with revision of the femoral stem or of any component as endpoints. At the time of this review, 76 patients without stem revision were assessed clinically and radiologically (mean follow-up 24.0 years (21.5 to 26.5)). For the 76 unrevised hips the mean Harris hip score was 87.1 (65 to 97). Femoral osteolysis was detected in five hips (6.6%) only in Gruen zone 7. Undersized stems were at higher risk of revision owing to aseptic loosening (p = 0.0003). Patients implanted with the stem in a varus position were at higher risk of femoral cortical hypertrophy and thigh pain (p = 0.0006 and p = 0.0007, respectively). In our study, survival, clinical outcome and radiographic data remained excellent in the third decade after implantation. Nonetheless, undersized stems were at higher risk of revision owing to aseptic loosening.

Steroid injections in the treatment of humeral unicameral bone cysts: long-term follow-up and review of the literature.

BACKGROUND: Retrospective evaluation of long-term effectiveness of the steroid injections treatment in patients with unicameral bone cysts (UBC). METHODS: From January 1993 to April 2005, 23 children affected by proximal humeral UBC were evaluated according to the Neer-Cole classification system and treated with serial methylprednisolone acetate's injections. The patients were followed up at 1, 3, 6 and 12 months and then every year until the adolescence. RESULTS: After treatment, in 15 out of 23 patients (65.2%), the humeral cysts were referred, respectively, as Grade 1 and in four as Grade 2. In 4 patients, a refracture occurred. Statistical analysis showed an overall good response in 82.6% of patients at the end of the follow-up. Minor complication including skin discoloration accounted for 13.04%. CONCLUSIONS: The steroid injections showed to be an alternative excellent treatment for UBC, with complete healing of the lesions in the majority of cases. This procedure is not expensive, mini-invasive, with low surgical risk and short hospitalization.

Congenital idiopathic talipes equinovarus: an evaluation in infants treated by the Ponseti method.

BACKGROUND: Congenital talipes equinovarus (CTEV) is a common but still not fully understood disorder of the lower limb. It is usually defined as a fixation of the foot in adduction, supination, and varus. Different treatment options exist including the Ponseti method. AIM: We report here the results obtained in infants with CTEV treated by the Ponseti method. PATIENTS AND METHODS: Eighty two patients (114 clubfeet) were enrolled at the Orthopaedic Clinic of Catania University during the period of March 2004 to January 2010 and followed prospectively up to February 2011: 56 patients (68.29%) were male, the anomaly was bilateral in 32 (39%) cases, unilateral in 50 (60.9%) in the right side in 28 (56%). The mean age at initiation of treatment was 14 days (range 3-81 days), severity of the club foot deformity by the Pirani Severity score was 5.56 points (range 4.3-6 points). Total numbers of Ponseti casts before tenotomy, details of tenotomy, and compliance with CTEV brace were recorded. Clinical evaluation was performed using the functional Ponseti Scoring System. Mean follow up was 4 years: range 13-83 months. RESULTS: An average of 6.6 casts was necessary before performing the tenotomy. Tenotomy was performed by a single surgeon (V.P.) in a total of 68 patients (82.93%) always in an operating room under general anaesthesia by a percutaneous approach at a mean age of 106 days (range 45-213 days). Compliance with CTEV brace was satisfactory in 79 patients (96.3%). Functional Ponseti Scores were good/excellent in 79 (96.34%) patients (109 clubfeet; 95.61%). Only 3 patients; 3.7% (5 clubfeet; 4.4%) suffered relapse. Poor compliance with the Denis Browne splint was thought to be the main cause of failure. CONCLUSIONS: The Ponseti method provides an excellent outcome at follow up in the treatment of congenital idiopathic clubfoot.

TGF-ß1 exposure induces epithelial to mesenchymal transition both in CSCs and non-CSCs of the A549 cell line, leading to an increase of migration ability in the CD133+ A549 cell fraction.

Metastasis is the leading cause of death by cancer. Non-small-cell lung cancer (NSCLC) represents nearly 85% of primary malignant lung tumours. Recent researches have demonstrated that epithelial-to-mesenchymal transition (EMT) plays a key role in the early process of metastasis of cancer cells. Transforming growth factor-ß1 (TGF-ß1) is the major inductor of EMT. The aim of this study is to investigate TGF-ß1's effect on cancer stem cells (CSCs) identified as cells positive for CD133, side population (SP) and non-cancer stem cells (non-CSCs) identified as cells negative for CD133, and SP in the A549 cell line. We demonstrate that TGF-ß1 induces EMT in both CSC and non-CSC A549 sublines, upregulating the expression of mesenchymal markers such as vimentin and Slug, and downregulating levels of epithelial markers such as e-cadherin and cytokeratins. CSC and non-CSC A549 sublines undergoing EMT show a strong migration and strong levels of MMP9 except for the CD133(-) cell fraction. OCT4 levels are strongly upregulated in all cell fractions except CD133(-) cells. On the contrary, wound size reveals that TGF-ß1 enhances motility in wild-type A549 as well as CD133(+) and SP(+) cells. For CD133(-) and SP(-) cells, TGF-ß1 exposure does not change the motility. Finally, assessment of growth kinetics reveals major colony-forming efficiency in CD133(+) A549 cells. In particular, SP(+) and SP(-) A549 cells show more efficiency to form colonies than untreated corresponding cells, while for CD133(-) cells no change in colony number was observable after TGF-ß1 exposure. We conclude that it is possible to highlight different cell subpopulations with different grades of stemness. Each population seems to be involved in different biological mechanisms such as stemness maintenance, tumorigenicity, invasion and migration.

Colonization and movement of GFP-labeled Clavibacter michiganensis subsp. michiganensis during tomato infection.

The vascular pathogen Clavibacter michiganensis subsp. michiganensis is responsible for bacterial wilt and canker of tomato. Pathogenicity of this bacterium is dependent on plasmid-borne virulence factors and serine proteases located on the chromosomal chp/tomA pathogenicity island (PAI). In this study, colonization patterns and movement of C. michiganensis subsp. michiganensis during tomato infection was examined using a green fluorescent protein (GFP)-labeled strain. A plasmid expressing GFP in C. michiganensis subsp. michiganensis was constructed and found to be stable in planta for at least 1 month. Confocal laser-scanning microscopy (CLSM) of inoculated stems showed that the pathogen extensively colonizes the lumen of xylem vessels and preferentially attaches to spiral secondary wall thickening of the protoxylem. Acropetal movement of the wild-type strain C. michiganensis subsp. michiganensis NCPPB382 (Cmm382) in tomato resulted in an extensive systemic colonization of the whole plant reaching the apical region after 15 days, whereas Cmm100 (lacking the plasmids pCM1 and pCM2) or Cmm27 (lacking the chp/tomA PAI) remained confined to the area surrounding of the inoculation site. Cmm382 formed biofilm-like structures composed of large bacterial aggregates on the interior of xylem walls as observed by CLSM and scanning electron microscopy. These findings suggest that virulence factors located on the chp/tomA PAI or the plasmids are required for effective movement of the pathogen in tomato and for the formation of cellular aggregates.

Plant adaptation to dynamically changing environment: the shade avoidance response.

The success of competitive interactions between plants determines the chance of survival of individuals and eventually of whole plant species. Shade-tolerant plants have adapted their photosynthesis to function optimally under low-light conditions. These plants are therefore capable of long-term survival under a canopy shade. In contrast, shade-avoiding plants adapt their growth to perceive maximum sunlight and therefore rapidly dominate gaps in a canopy. Daylight contains roughly equal proportions of red and far-red light, but within vegetation that ratio is lowered as a result of red absorption by photosynthetic pigments. This light quality change is perceived through the phytochrome system as an unambiguous signal of the proximity of neighbors resulting in a suite of developmental responses (termed the shade avoidance response) that, when successful, result in the overgrowth of those neighbors. Shoot elongation induced by low red/far-red light may confer high relative fitness in natural dense communities. However, since elongation is often achieved at the expense of leaf and root growth, shade avoidance may lead to reduction in crop plant productivity. Over the past decade, major progresses have been achieved in the understanding of the molecular basis of shade avoidance. However, uncovering the mechanisms underpinning plant response and adaptation to changes in the ratio of red to far-red light is key to design new strategies to precise modulate shade avoidance in time and space without impairing the overall crop ability to compete for light.

Ganglioglioma of the right lateral ventricle approached with neuronavigation and intraoperative DTI. Case report and literature review.

UNLABELLED: BACKGROUND AND STUDY OBJECT: We describe the case of a young patient treated for a ganglioglioma of the right lateral ventricle. The ganglioma was approached via a temporal route using neuronavigation. Such lesions, if confined to the ventricular system and, especially, to the lateral ventricle, are unusual. CASE REPORT: A 22-year-old male was admitted because of headache and seizures. Magnetic resonance imaging (MRI) revealed a neoplasm in the temporal horn of the right lateral ventricle. A CT scan demonstrated calcifications. After preoperative planning, the patient was scheduled for neuronavigated operation. A small temporal craniotomy was performed. Once the dura was opened, we acquired a volumetric intraoperative MRI with diffusion tensor imaging (DTI). The optic tract was visualized again on intraoperative DTI and the volumetric MRI was used to update the navigational data. RESULTS: The temporal corticotomy between the inferior and the middle temporal gyrus was guided by neuronavigation, keeping the surgical corridor in distance to the optic tract until the tumor was reached. After tumor resection, we repeated intraoperative MRI with DTI, which confirmed complete tumor removal and the patency of the optic pathway. Our protocol allowed us to reach the lesion via a minimally invasive approach with near real-time control of the integrity of the optic tract during surgery. DTI was acquired preoperatively and during surgery, avoiding inaccuracy due to brain shift. CONCLUSIONS: Intraoperative MRI with DTI allows to treat such lesions via minimally invasive approaches and to reduce operative morbidity due to fiber tract damage.

Sequential expression of bacterial virulence and plant defense genes during infection of tomato with Clavibacter michiganensis subsp. michiganensis.

The molecular interactions between Clavibacter michiganensis subsp. michiganensis and tomato plant were studied by following the expression of bacterial virulence and host-defense genes during early stages of infection. The C. michiganensis subsp. michiganensis genes included the plasmid-borne cellulase (celA) and the serine protease (pat-1), and the serine proteases chpC and ppaA, residing on the chp/tomA pathogenicity island (PAI). Gene expression was measured following tomato inoculation with Cmm382 (wild type), Cmm100 (lacking the plasmids pCM1 and pCM2), and Cmm27 (lacking the PAI). Transcriptional analysis revealed that celA and pat-1 were significantly induced in Cmm382 at initial 12 to 72 h, whereas chpC and ppaA were highly expressed only 96 h after inoculation. Interdependence between the expression of chromosomal and of plasmid-located genes was revealed: expression of celA and pat-1 was substantially reduced in the absence of the chp/tomA PAI, whereas chpC and ppaA expressions were reduced in the absence of the virulence plasmids. Transcription of chromosomal genes involved in cell wall degradation (i.e., pelA1, celB, xysA, and xysB), was also induced at early stages of infection. Expression of the host-defense genes, chitinase class II and pathogenesis-related protein-5 isoform was induced in the absence of the PAI at early stages of infection, suggesting that PAI-located genes are involved in suppression of tomato basal defenses.

Riluzole restores motor activity in rats with post-traumatic peripheral neuropathy.

Riluzole is a presynaptic inhibitor of glutamate release with neuroprotective properties. In order to evaluate the effects of riluzole on motor activity in post-traumatic peripheral neuropathy (PTPN), the sciatic nerve of Wistar male rats was exposed monolaterally and subjected to crushing for one min by a surgical forceps. Animals received an intraperitoneal treatment with riluzole (2, 4 or 8 mg/kg per day), diclofenac (5, 10 or 20 mg/kg) or with vehicle for 3 days. Motor activity and coordination was evaluated in a circular open field and in the rotorod test. The treatment with riluzole stimulated ambulation in PTPN rats and improved their motor performance and coordination. The effect of treatment with riluzole on locomotor activity was greater than that of treatment with diclofenac and was dose-dependent. Furthermore, in contrast to vehicle- and diclofenac-treated rats, animals treated with riluzole showed a long-lasting improvement of locomotor activity as it was assessed 7 days after the end of treatment. These findings suggest that riluzole may improve motor performance in PTPN, and this does not depend on its antinociceptive activity. Its neuroprotective properties are possibly involved in this effect.

Heterotopic ossification after total hip replacement and the HLA system in the Sicilian population.

Heterotopic ossification (HO) is a frequent complication following total hip arthroplasty (THA). At present, the etiology HO is unknown, however, genetic predisposition may be a cause of HO in individuals in whom no risk factors can be detected. The goal of this study was to investigate the HLA system, searching for any correlation with the presence of HO after THA. Thirty-five patients of Sicilian origin were operated on between January 1997 and January 1999 for cementless THA under regional anesthesia. The entire series was divided into three groups and all underwent histocompatibility typing. Group I was made up of 10 patients who presented with HO Brooker grades 1 and 2 after THA; group 2 comprised 7 patients affected by grades 3 and 4 HO after THA; and group 3 was made up of 18 subjects who presented with one or more preoperative risk factors for developing peri-prosthetic HO before undergoing THA. No positivity for HLA-B27 antigen was observed, but there was as an increase in HLA-B18 (with respect to that in the Sicilian population) in patients with HO following THA. The main conclusion from the study is that there is a strong correlation between the presence of the antigens HLA-A2 and HLA-B18 in patients with HO grades 3 and 4.

Innate immunity in plants.

Studies of receptors and signal-transduction components that play a role in plant disease resistance have revealed remarkable similarities with innate immunity pathways in insects and mammals. In plants, specific receptors encoded by disease-resistance genes interact with products of microbial effector genes to activate defence responses. Resistance proteins have been found to have motifs in common with components of immune response pathways in mammals and invertebrates, and to rely on similar downstream signalling components. In the future, the sharing of ideas among plant and animal biologists is likely to broaden our understanding of defence responses in diverse organisms.

Signal recognition and transduction mediated by the tomato Pto kinase: a paradigm of innate immunity in plants.

Plant disease resistance is the result of an innate host defense mechanism, which relies on the ability of the plant to recognize pathogen invasion and to efficiently mount defense responses. In tomato, resistance to the pathogen Pseudomonas syringae is mediated by the specific interaction between the plant serine/threonine kinase Pto and the bacterial protein AvrPto. This article reviews molecular and biochemical properties that confer to Pto the capability to function as an intracellular receptor and to activate a signaling cascade leading to the induction of defense responses.

Thr38 and Ser198 are Pto autophosphorylation sites required for the AvrPto-Pto-mediated hypersensitive response.

The tomato Pto kinase confers resistance to Pseudomonas syringae pv. tomato expressing the AvrPto protein. To elucidate the role of Pto autophosphorylation in disease resistance, eight sites autophosphorylated by Pto in vitro were identified by a combination of HPLC purification of tryptic phosphopeptides, MALDI-TOF/MS analysis and Edman degradation. Mutational analysis of the autophosphorylation sites revealed that Pto residues Thr38 and Ser198 are required for AvrPto-Pto- mediated elicitation of a hypersensitive response in the plant. Thr38, which is the main Pto autophosporylation site and is located outside the kinase catalytic domain, was also required for Pto kinase activity and its physical interaction with AvrPto, the Pti1 kinase and the transcription factor Pti4. Ser198, located in the Pto activation domain, was dispensable for kinase activity and for interaction with AvrPto. However, a mutation at this site resulted in altered Pto interactions with the Pti1 kinase and the Pto interactors of unknown function Pti3 and Pti10. These results suggest that autophosphorylation events at Pto Thr38 and Ser198 are required for signal transduction by Pto and participate in distinct molecular mechanisms.

The ethylene-inducible PK12 kinase mediates the phosphorylation of SR splicing factors.

The tobacco PK12 is induced by the plant hormone ethylene and is a member of the LAMMER family of protein kinases. Members of this family contain in their C-terminus a unique 'EHLAMMERI/VLGPLP' motif of unknown function, and are related to cyclin- and mitogen-activated protein (MAP)-dependent kinases. The animal members of this class play a role in differentiation. They phosphorylate and physically interact with serine/arginine-rich (SR) splicing factors in vivo to alter their activity and the splicing of target mRNAs. SR proteins have been recently described in plants. The capability of PK12 LAMMER kinase to bind and phosphorylate SR proteins was tested in vitro by kinase and binding assays. The tobacco PK12 phosphorylated both animal and plant SR proteins and specifically interacted with the plant splicing factor atSRp34/SR1. In addition, by site-directed mutagenesis, the LAMMER motif was found to be required for PK12 kinase activity but was not necessary for substrate binding. Consistent with a role in phosphorylation of splicing factors, PK12 was found to localize to the nucleus when transiently over-expressed in suspension cells.

Subjects positive for hepatitis C virus RNA with normal aminotransferase levels, a "trompe l'oeil" clinical picture?

BACKGROUND: Quite often subjects affected by chronic hepatitis C virus infection have no clinical signs of liver disease and serum aminotransferase values never go beyond the upper limit of normal. Yet these subjects, defined "asymptomatic HCV carriers", often have active viral replication and various degrees of histological damage. AIMS: To verify, in a population of antibody to hepatitis C virus carriers, if normal serum aminotransferase values in hepatitis C virus-RNA positive differed considerably from those in hepatitis C virus-RNA negative subjects. SUBJECTS/METHODS: We followed 24 anti-hepatitis C virus-positive subjects (15 hepatitis C virus-RNA positive and 9 negative) by measuring alanine and aspartate aminotransferase levels at 3-month intervals for a median of 40 months (range 6-77). RESULTS: Determinations resulted repeatedly and rigorously within the normal range in all participants. Alanine aminotransferase values were higher in hepatitis C virus-RNA positives than in negatives (mean +/- SD: 0.609+/-0.172 vs 0.434+/-0.153 times the upper limit of normal; p

The major site of the pti1 kinase phosphorylated by the pto kinase is located in the activation domain and is required for pto-pti1 physical interaction.

The Pto and Pti1 serine/threonine protein kinases are key components of the signaling pathway leading to speck disease resistance in tomato. The two kinases physically interact in the yeast two-hybrid system, and Pto specifically phosphorylates Pti1 in vitro. In this study, we identified and characterized the major Pti1 site phosphorylated by Pto. Pto was expressed in Escherichia coli as a maltose-binding fusion protein (MBP-Pto), and used to phosphorylate in vitro a kinase deficient Pti1 protein fused to glutathione S-transferase (GST-Pti1[K96N]). The major phosphopeptide derived from trypsin digestion of phosphorylated GST-Pti1(K96N) was partially purified by reverse-phase HPLC and analyzed by matrix assisted laser desorption/ionization mass spectrometry. Its mass corresponded to phosphopeptide LHSTR, which lies in the Pti1 kinase activation domain at amino acid position 230-234. By phosphoamino acid analysis, Thr233 was determined to be the phosphorylation site of peptide LHSTR. Mutations of Thr233 reduced dramatically Pti1 phosphorylation by MBP-Pto and Pti1 autophosphorylation, providing evidence that the same Pti1 site is involved in the two reactions. Moreover, phosphorylation of Thr233 appeared to be required for Pto-Pti1 physical interaction, as a mutation of this site to alanine, but not to aspartate, abolished the interaction between Pto and Pti1 in the yeast two-hybrid system.