CHEMOSENSITIZATION OF HEPATOCELLULAR CARCINOMA TO GEMCITABINE BY NON-INVASIVE RADIOFREQUENCY FIELD-INDUCED HYPERTHERMIA

Gemcitabine is a potent nucleoside analogue against solid tumors however drug resistance rapidly emerges. Removal of gemcitabine incorporated in the DNA by repair mechanisms could potentially contribute to resistance in chemo-refractory solid tumors. In this study, we evaluated homologous recombination repair of gemcitabine-stalled replication forks as a potential mechanism contributing to resistance. We also studied the effect of hyperthermia on homologous recombination pathway to explain the previously reported synergy between gemcitabine and hyperthermia. We found that hyperthermia degrades and inhibits localization of Mre11 to gemcitabine-stalled replication forks. Furthermore, gemcitabine-treated cells that were also treated with hyperthermia demonstrate a prolonged passage through late S/ G2 phase of cell cycle in comparison to cells treated with gemcitabine alone. This coincides with inhibition of resolution of γH2AX foci. Our findings also demonstrate that thermal sensitization of human hepatocellular carcinoma cell lines to gemcitabine is mediated through an Mre11-dependent homologous recombination repair pathway. Combination of non-invasive radiofrequency field-induced hyperthermia and gemcitabine was superior to either therapy alone (p

Genetic Predictors of Clinical Outcomes in Non-Small Cell Lung Cancer Patients

Lung cancer is the leading cause of cancer-related mortality in the US. Emerging evidence has shown that host genetic factors can interact with environmental exposures to influence patient susceptibility to the diseases as well as clinical outcomes, such as survival and recurrence. We aimed to identify genetic prognostic markers for non-small cell lung cancer (NSCLC), a major (85%) subtype of lung cancer, and also in other subgroups. With the fast evolution of genotyping technology, genetic association studies have went through candidate gene approach, to pathway-based approach, to the genome wide association study (GWAS). Even in the era of GWAS, pathway-based approach has its own advantages on studying cancer clinical outcomes: it is cost-effective, requiring a smaller sample size than GWAS easier to identify a validation population and explore gene-gene interactions. In the current study, we adopted pathway-based approach focusing on two critical pathways - miRNA and inflammation pathways. MicroRNAs (miRNA) post-transcriptionally regulate around 30% of human genes. Polymorphisms within miRNA processing pathways and binding sites may influence patients’ prognosis through altered gene regulation. Inflammation plays an important role in cancer initiation and progression, and also has shown to impact patients’ clinical outcomes. We first evaluated 240 single nucleotide polymorphisms (SNPs) in miRNA biogenesis genes and predicted binding sites in NSCLC patients to determine associations with clinical outcomes in early-stage (stage I and II) and late-stage (stage III and IV) lung cancer patients, respectively. First, in 535 early-stage patients, after correcting multiple comparisons, FZD4:rs713065 (hazard ratio [HR]:0.46, 95% confidence interval [CI]:0.32-0.65) showed a significant inverse association with survival in early stage surgery-only patients. SP1:rs17695156 (HR:2.22, 95% CI:1.44-3.41) and DROSHA:rs6886834 (HR:6.38, 95% CI:2.49-16.31) conferred increased risk of progression in the all patients and surgery-only populations, respectively. FAS:rs2234978 was significantly associated with improved survival in all patients (HR:0.59, 95% CI:0.44-0.77) and in the surgery plus chemotherapy populations (HR:0.19, 95% CI:0.07-0.46).. Functional genomics analysis demonstrated that this variant creates a miR-651 binding site resulting in altered miRNA regulation of FAS, providing biological plausibility for the observed association. We then analyzed these associations in 598 late-stage patients. After multiple comparison corrections, no SNPs remained significant in the late stage group, while the top SNP NAT1:rs15561 (HR=1.98, 96%CI=1.32-2.94) conferred a significantly increased risk of death in the chemotherapy subgroup. To test the hypothesis that genetic variants in the inflammation-related pathways may be associated with survival in NSCLC patients, we first conducted a three-stage study. In the discovery phase, we investigated a comprehensive panel of 11,930 inflammation-related SNPs in three independent lung cancer populations. A missense SNP (rs2071554) in HLA-DOB was significantly associated with poor survival in the discovery population (HR: 1.46, 95% CI: 1.02-2.09), internal validation population (HR: 1.51, 95% CI: 1.02-2.25), and external validation (HR: 1.52, 95% CI: 1.01-2.29) population. Rs2900420 in KLRK1 was significantly associated with a reduced risk for death in the discovery (HR: 0.76, 95% CI: 0.60-0.96) and internal validation (HR: 0.77, 95% CI: 0.61-0.99) populations, and the association reached borderline significance in the external validation population (HR: 0.80, 95% CI: 0.63-1.02). We also evaluated these inflammation-related SNPs in NSCLC patients in never smokers. Lung cancer in never smokers has been increasingly recognized as distinct disease from that in ever-smokers. A two-stage study was performed using a discovery population from MD Anderson (411 patients) and a validation population from Mayo Clinic (311 patients). Three SNPs (IL17RA:rs879576, BMP8A:rs698141, and STK:rs290229) that were significantly associated with survival were validated (pCD74:rs1056400 and CD38:rs10805347) were borderline significant (p=0.08) in the Mayo Clinic population. In the combined analysis, IL17RA:rs879576 resulted in a 40% reduction in the risk for death (p=4.1 × 10-5 [p=0.61, heterogeneity test]). We also validated a survival tree created in MD Anderson population in the Mayo Clinic population. In conclusion, our results provided strong evidence that genetic variations in specific pathways that examined (miRNA and inflammation pathways) influenced clinical outcomes in NSCLC patients, and with further functional studies, the novel loci have potential to be translated into clinical use.

Exposure to nonsteroidal anti-inflammatory drugs and weight loss and hospitalization in non-small cell lung cancer patients

Background. Cancer cachexia is a common syndrome complex in cancer, occurring in nearly 80% of patients with advanced cancer and responsible for at least 20% of all cancer deaths. Cachexia is due to increased resting energy expenditure, increased production of inflammatory mediators, and changes in lipid and protein metabolism. Non-steroidal anti-inflammatory drugs (NSAIDs), by virtue of their anti-inflammatory properties, are possibly protective against cancer-related cachexia. Since cachexia is also associated with increased hospitalizations, this outcome may also show improvement with NSAID exposure. ^ Design. In this retrospective study, computerized records from 700 non-small cell lung cancer patients (NSCLC) were reviewed, and 487 (69.57%) were included in the final analyses. Exclusion criteria were severe chronic obstructive pulmonary disease, significant peripheral edema, class III or IV congestive heart failure, liver failure, other reasons for weight loss, or use of research or anabolic medications. Information on medication history, body weight and hospitalizations was collected from one year pre-diagnosis until three years post-diagnosis. Exposure to NSAIDs was defined if a patient had a history of being treated with NSAIDs for at least 50% of any given year in the observation period. We used t-test and chi-square tests for statistical analyses. ^ Results. Neither the proportion of patients with cachexia (p=0.27) nor the number of hospitalizations (p=0.74) differed among those with a history of NSAID use (n=92) and those without (n=395). ^ Conclusions. In this study, NSAID exposure was not significantly associated with weight loss or hospital admissions in patients with NSCLC. Further studies may be needed to confirm these observations.^

Post-transcriptional Regulation of Mammalian Gene Expression in Non-coding Region of Target RNA

Tumor Suppressor Candidate 2 (TUSC2) is a novel tumor suppressor gene located in the human chromosome 3p21.3 region. TUSC2 mRNA transcripts could be detected on Northern blots in both normal lung and some lung cancer cell lines, but no endogenous TUSC2 protein could be detected in a majority of lung cancer cell lines. Mechanisms regulating TUSC2 protein expression and its inactivation in primary lung cancer cells are largely unknown. We investigated the role of the 5’- and 3’-untranslated regions (UTRs) of the TUSC2 gene in the regulation of TUSC2 protein expression. We found that two small upstream open-reading frames (uORFs) in the 5’UTR of TUSC2 could markedly inhibit the translational initiation of TUSC2 protein by interfering with the “scanning” of the ribosome initiation complexes. Site-specific stem-loop array reverse transcription-polymerase chain reaction (SLA-RT-PCR) verified several micoRNAs (miRNAs) targeted at 3’UTR and directed TUSC2 cleavage and degradation. In addition, we used the established let-7-targeted high mobility group A2 (Hmga2) mRNA as a model system to study the mechanism of regulation of target mRNA by miRNAs in mammalian cells under physiological conditions. There have been no evidence of direct link between mRNA downregulation and mRNA cleavages mediated by miRNAs. Here we showed that the endonucleolytic cleavages on mRNAs were initiated by mammalian miRNA in seed pairing style. Let-7 directed cleavage activities among the eight predicted potential target sites have varied efficiency, which are influenced by the positional and the structural contexts in the UTR. The 5’ cleaved RNA fragments were mostly oligouridylated at their 3’-termini and accumulated for delayed 5’–3’ degradation. RNA fragment oligouridylation played important roles in marking RNA fragments for delayed bulk degradation and in converting RNA degradation mode from 3’–5’ to 5’–3’ with cooperative efforts from both endonucleolytic and non-catalytic miRNA-induced silencing complex (miRISC). Our findings point to a mammalian miRNA-mediated mechanism for the regulation of mRNA that miRNA can decrease target mRNA through target mRNA cleavage and uridine addition

The role of Ski/Sno oncoproteins as negative regulators of the TGF-beta/SMAD signaling pathway in B -cell non -Hodgkin's lymphoma (NHL-B)

Non-Hodgkin's lymphomas are common tumors of the human immune system, primarily of B cell lineage (NHL-B). Negative growth regulation in the B cell lineage is mediated primarily through the TGF-β/SMAD signaling pathway that regulates a variety of tumor suppressor genes. Ski was originally identified as a transforming oncoprotein, whereas SnoN is an isoform of the Sno protein that shares a large region of homology with Ski. In this study, we show that Ski/SnoN are endogenously over-expressed both in patients' lymphoma cells and NHL-B cell lines. Exogenous TGF-β1 treatment induces down-regulation of Ski and SnoN oncoprotein expression in an NHL-B cell line, implying that Ski and SnoN modulate the TGF-β signaling pathway and are involved in cell growth regulation. Furthermore, we have developed an NHL-B cell line (DB) that has a null mutation in TGF-β receptor type II. In this mutant cell line, Ski/SnoN proteins are not down-regulated in response to TGF-β1 treatment, suggesting that downregulation of Ski and SnoN proteins in NHL-B require an intact functional TGF-β signaling pathway Resting normal B cells do not express Ski until activated by antigens and exogenous cytokines, whereas a low level of SnoN is also present in peripheral blood Go B cells. In contrast, autonomously growing NHL-B cells over-express Ski and SnoN, implying that Ski and SnoN are important cell cycle regulators. To further investigate a possible link between reduction of the Ski protein level and growth inhibition, Ski antisense oligodeoxynucleotides were transfected into NHL-B cells. The Ski protein level was found to decrease to less than 40%, resulting in restoring the effect of TGF-β and leading to cell growth inhibition and G1 cell cycle arrest. Co-immunoprecipitation experiments demonstrated that Ski associates with Smad4 in the nucleus, strongly suggesting that over-expression of the nuclear protein Ski and/or SnoN negatively regulates the TGF-β pathway, possibly by modulating Smad-mediated tumor suppressor gene expression. Together, in NHL-B, the TGF-β/SMAD growth inhibitory pathway is usually intact, but over-expression of the Ski and/or SnoN, which binds to Smad4, abrogates the negative regulatory effects of TGF-β/SMAD in lymphoma cell growth and potentiates the growth potential of neoplastic B cells. ^

Dysregulated CD40-NF-kappaB signaling in the pathophysiology of aggressive non -Hodgkin's lymphoma B cells

Non-Hodgkin's Lymphomas (NHL) are a group (>30) of important human lymphoid cancers that unlike other tumors today, are showing a marked increase in incidence. The lack of insight to the pathogenesis of B-cell NHL poses a significant problem in the early detection and effective treatment of these malignancies. This study shows that large B-cell lymphoma (LBCL) cells, the most common type of B-cell NHL (account for more than 30% of cases), have developed a novel mechanism for autonomous neoplastic B cell growth. We have identified that the key transcription factor NF-κB, is constitutively activated in LBCL cell lines and primary biopsy-derived LBCL cells, suggesting that they are autonomously activated, and do not require accessory T-cell signaling for cell growth and survival. Further studies have indicated that LBCL cells ectopically express an important T-cell associated co-mitogenic factor, CD154 (CD40 ligand), that is able to internally activate the CD401NF-κB pathway, through constitutive binding to its cognate receptor, CD40, on the lymphoma cell surface. CD40 activation triggers the formation of a “Signalosome” comprising virtually the entire canonical CD40/NF-κB signaling pathway that is anchored by CD40 in plasma membrane lipid rafts. The CD40 Signalosome is vulnerable to interdiction by antibody against CD40 that disrupts the Signalosome and induces cell death in the malignant cells. In addition to constitutive NF-κB activation, we have found that the nuclear factor of activated T cells (NFAT) transcription factor is also constitutively activated in LBCL cells. We have demonstrated that the constitutively active NFATc1 and c-rel members of the NFAT and NF-κB families of transcription factors, respectively, interact with each other, bind to the CD154 promoter, and synergistically activate CD154 gene transcription. Down-regulation of NFATc1 and c-rel with small interfering RNA inhibits CD154 gene transcription and lymphoma cell growth. Our findings suggest that continuous CD40 activation not only provides dysregulated proliferative stimuli for lymphoma cell growth and extended tumor cell survival, but also allows continuous regeneration of the CD40 ligand in the lymphoma cell and thereby recharges the system through a positive feedback mechanism. Targeting the CD40/NF-κB signaling pathway could provide potential therapeutic modalities for LBCL cells in the future. ^

DBpedia spotlight: shedding light on the web of documents

Interlinking text documents with Linked Open Data enables the Web of Data to be used as background knowledge within document-oriented applications such as search and faceted browsing. As a step towards interconnecting the Web of Documents with the Web of Data, we developed DBpedia Spotlight, a system for automatically annotating text documents with DBpedia URIs. DBpedia Spotlight allows users to congure the annotations to their specic needs through the DBpedia Ontology and quality measures such as prominence, topical pertinence, contextual ambiguity and disambiguation condence. We compare our approach with the state of the art in disambiguation, and evaluate our results in light of three baselines and six publicly available annotation systems, demonstrating the competitiveness of our system. DBpedia Spotlight is shared as open source and deployed as a Web Service freely available for public use.

DBpedia Spotlight: Shedding Light on the Web of Documents

Interlinking text documents with Linked Open Data enables the Web of Data to be used as background knowledge within document-oriented applications such as search and faceted browsing. As a step towards interconnecting the Web of Documents with the Web of Data, we developed DBpedia Spotlight, a system for automatically annotating text documents with DBpedia URIs. DBpedia Spotlight allows users to configure the annotations to their specific needs through the DBpedia Ontology and quality measures such as prominence, topical pertinence, contextual ambiguity and disambiguation confidence. We compare our approach with the state of the art in disambiguation, and evaluate our results in light of three baselines and six publicly available annotation systems, demonstrating the competitiveness of our system. DBpedia Spotlight is shared as open source and deployed as a Web Service freely available for public use.

A Method for Developing Ontologies from User-Generated Classication Systems

Abstract Web 2.0 applications enabled users to classify information resources using their own vocabularies. The bottom-up nature of these user-generated classification systems have turned them into interesting knowledge sources, since they provide a rich terminology generated by potentially large user communities. Previous research has shown that it is possible to elicit some emergent semantics from the aggregation of individual classifications in these systems. However the generation of ontologies from them is still an open research problem. In this thesis we address the problem of how to tap into user-generated classification systems for building domain ontologies. Our objective is to design a method to develop domain ontologies from user-generated classifications systems. To do so, we rely on ontologies in the Web of Data to formalize the semantics of the knowledge collected from the classification system. Current ontology development methodologies have recognized the importance of reusing knowledge from existing resources. Thus, our work is framed within the NeOn methodology scenario for building ontologies by reusing and reengineering non-ontological resources. The main contributions of this work are: An integrated method to develop ontologies from user-generated classification systems. With this method we extract a domain terminology from the classification system and then we formalize the semantics of this terminology by reusing ontologies in the Web of Data. Identification and adaptation of existing techniques for implementing the activities in the method so that they can fulfill the requirements of each activity. A novel study about emerging semantics in user-generated lists. Resumen La web 2.0 permitió a los usuarios clasificar recursos de información usando su propio vocabulario. Estos sistemas de clasificación generados por usuarios son recursos interesantes para la extracción de conocimiento debido principalmente a que proveen una extensa terminología generada por grandes comunidades de usuarios. Se ha demostrado en investigaciones previas que es posible obtener una semántica emergente de estos sistemas. Sin embargo la generación de ontologías a partir de ellos es todavía un problema de investigación abierto. Esta tesis trata el problema de cómo aprovechar los sistemas de clasificación generados por usuarios en la construcción de ontologías de dominio. Así el objetivo de la tesis es diseñar un método para desarrollar ontologías de dominio a partir de sistemas de clasificación generados por usuarios. El método propuesto reutiliza conceptualizaciones existentes en ontologías publicadas en la Web de Datos para formalizar la semántica del conocimiento que se extrae del sistema de clasificación. Por tanto, este trabajo está enmarcado dentro del escenario para desarrollar ontologías mediante la reutilización y reingeniería de recursos no ontológicos que se ha definido en la Metodología NeOn. Las principales contribuciones de este trabajo son: Un método integrado para desarrollar una ontología de dominio a partir de sistemas de clasificación generados por usuarios. En este método se extrae una terminología de dominio del sistema de clasificación y posteriormente se formaliza su semántica reutilizando ontologías en la Web de Datos. La identificación y adaptación de un conjunto de técnicas para implementar las actividades propuestas en el método de tal manera que puedan cumplir automáticamente los requerimientos de cada actividad. Un novedoso estudio acerca de la semántica emergente en las listas generadas por usuarios en la Web.

Cross-lingual ontology matching as a challenge for the Multilingual Semantic Web

Recently, the Semantic Web has experienced significant advancements in standards and techniques, as well as in the amount of semantic information available online. Nevertheless, mechanisms are still needed to automatically reconcile information when it is expressed in different natural languages on the Web of Data, in order to improve the access to semantic information across language barriers. In this context several challenges arise [1], such as: (i) ontology translation/localization, (ii) cross-lingual ontology mappings, (iii) representation of multilingual lexical information, and (iv) cross-lingual access and querying of linked data. In the following we will focus on the second challenge, which is the necessity of establishing, representing and storing cross-lingual links among semantic information on the Web. In fact, in a “truly” multilingual Semantic Web, semantic data with lexical representations in one natural language would be mapped to equivalent or related information in other languages, thus making navigation across multilingual information possible for software agents.

Quality of Service (QoS) in MPLS networks

Telecommunications networks have been always expanding and thanks to it, new services have appeared. The old mechanisms for carrying packets have become obsolete due to the new service requirements, which have begun working in real time. Real time traffic requires strict service guarantees. When this traffic is sent through the network, enough resources must be given in order to avoid delays and information losses. When browsing through the Internet and requesting web pages, data must be sent from a server to the user. If during the transmission there is any packet drop, the packet is sent again. For the end user, it does not matter if the webpage loads in one or two seconds more. But if the user is maintaining a conversation with a VoIP program, such as Skype, one or two seconds of delay in the conversation may be catastrophic, and none of them can understand the other. In order to provide support for this new services, the networks have to evolve. For this purpose MPLS and QoS were developed. MPLS is a packet carrying mechanism used in high performance telecommunication networks which directs and carries data using pre-established paths. Now, packets are forwarded on the basis of labels, making this process faster than routing the packets with the IP addresses. MPLS also supports Traffic Engineering (TE). This refers to the process of selecting the best paths for data traffic in order to balance the traffic load between the different links. In a network with multiple paths, routing algorithms calculate the shortest one, and most of the times all traffic is directed through it, causing overload and packet drops, without distributing the packets in the other paths that the network offers and do not have any traffic. But this is not enough in order to provide the real time traffic the guarantees it needs. In fact, those mechanisms improve the network, but they do not make changes in how the traffic is treated. That is why Quality of Service (QoS) was developed. Quality of service is the ability to provide different priority to different applications, users, or data flows, or to guarantee a certain level of performance to a data flow. Traffic is distributed into different classes and each of them is treated differently, according to its Service Level Agreement (SLA). Traffic with the highest priority will have the preference over lower classes, but this does not mean it will monopolize all the resources. In order to achieve this goal, a set policies are defined to control and alter how the traffic flows. Possibilities are endless, and it depends in how the network must be structured. By using those mechanisms it is possible to provide the necessary guarantees to the real-time traffic, distributing it between categories inside the network and offering the best service for both real time data and non real time data. Las Redes de Telecomunicaciones siempre han estado en expansión y han propiciado la aparición de nuevos servicios. Los viejos mecanismos para transportar paquetes se han quedado obsoletos debido a las exigencias de los nuevos servicios, que han comenzado a operar en tiempo real. El tráfico en tiempo real requiere de unas estrictas garantías de servicio. Cuando este tráfico se envía a través de la red, necesita disponer de suficientes recursos para evitar retrasos y pérdidas de información. Cuando se navega por la red y se solicitan páginas web, los datos viajan desde un servidor hasta el usuario. Si durante la transmisión se pierde algún paquete, éste se vuelve a mandar de nuevo. Para el usuario final, no importa si la página tarda uno o dos segundos más en cargar. Ahora bien, si el usuario está manteniendo una conversación usando algún programa de VoIP (como por ejemplo Skype) uno o dos segundos de retardo en la conversación podrían ser catastróficos, y ninguno de los interlocutores sería capaz de entender al otro. Para poder dar soporte a estos nuevos servicios, las redes deben evolucionar. Para este propósito se han concebido MPLS y QoS MPLS es un mecanismo de transporte de paquetes que se usa en redes de telecomunicaciones de alto rendimiento que dirige y transporta los datos de acuerdo a caminos preestablecidos. Ahora los paquetes se encaminan en función de unas etiquetas, lo cual hace que sea mucho más rápido que encaminar los paquetes usando las direcciones IP. MPLS también soporta Ingeniería de Tráfico (TE). Consiste en seleccionar los mejores caminos para el tráfico de datos con el objetivo de balancear la carga entre los diferentes enlaces. En una red con múltiples caminos, los algoritmos de enrutamiento actuales calculan el camino más corto, y muchas veces el tráfico se dirige sólo por éste, saturando el canal, mientras que otras rutas se quedan completamente desocupadas. Ahora bien, esto no es suficiente para ofrecer al tráfico en tiempo real las garantías que necesita. De hecho, estos mecanismos mejoran la red, pero no realizan cambios a la hora de tratar el tráfico. Por esto es por lo que se ha desarrollado el concepto de Calidad de Servicio (QoS). La calidad de servicio es la capacidad para ofrecer diferentes prioridades a las diferentes aplicaciones, usuarios o flujos de datos, y para garantizar un cierto nivel de rendimiento en un flujo de datos. El tráfico se distribuye en diferentes clases y cada una de ellas se trata de forma diferente, de acuerdo a las especificaciones que se indiquen en su Contrato de Tráfico (SLA). EL tráfico con mayor prioridad tendrá preferencia sobre el resto, pero esto no significa que acapare la totalidad de los recursos. Para poder alcanzar estos objetivos se definen una serie de políticas para controlar y alterar el comportamiento del tráfico. Las posibilidades son inmensas dependiendo de cómo se quiera estructurar la red. Usando estos mecanismos se pueden proporcionar las garantías necesarias al tráfico en tiempo real, distribuyéndolo en categorías dentro de la red y ofreciendo el mejor servicio posible tanto a los datos en tiempo real como a los que no lo son.

How flowering plants discriminate between self and non-self pollen to prevent inbreeding.

Flowering plants have evolved various genetic mechanisms to circumvent the tendency for self-fertilization created by the close proximity of male and female reproductive organs in a bisexual flower. One such mechanism is gametophytic self-incompatibility, which allows the female reproductive organ, the pistil, to distinguish between self pollen and non-self pollen; self pollen is rejected, whereas non-self pollen is accepted for fertilization. The Solanaceae family has been used as a model to study the molecular and biochemical basis of self/non-self-recognition and self-rejection. Discrimination of self and non-self pollen by the pistil is controlled by a single polymorphic locus, the S locus. The protein products of S alleles in the pistil, S proteins, were initially identified based on their cosegregation with S alleles. S proteins have recently been shown to indeed control the ability of the pistil to recognize and reject self pollen. S proteins are also RNases, and the RNase activity has been shown to be essential for rejection of self pollen, suggesting that the biochemical mechanism of self-rejection involves the cytotoxic action of the RNase activity. S proteins contain various numbers of N-linked glycans, but the carbohydrate moiety has been shown not to be required for the function of S proteins, suggesting that the S allele specificity determinant of S proteins lies in the amino acid sequence. The male component in self-incompatibility interactions, the pollen S gene, has not yet been identified. The possible nature of the pollen S gene product and the possible mechanism by which allele-specific rejection of pollen is accomplished are discussed.