Role In Intercellular Signaling Research Articles

Mesenchymal stem cell-based therapy for autoimmune diseases: emerging roles of extracellular vesicles

In autoimmune disease body's own immune system knows healthy cells as undesired and foreign cells. Over 80 types of autoimmune diseases have been recognized. Currently, at clinical practice, treatment strategies for autoimmune disorders are based on relieving symptoms and preventing difficulties. In other words, there is no effective and useful therapy up to now. It has been well-known that mesenchymal stem cells (MSCs) possess immunomodulatory effects. This strongly suggests that MSCs might be as a novel modality for treatment of autoimmune diseases. Supporting this notion a few preclinical and clinical studies indicate that MSCs ameliorate autoimmune disorders. Interestingly, it has been found that the beneficial effects of MSCs in autoimmune disorders are not relying only on direct cell-to-cell communication but on their capability to produce a broad range of paracrine factors including growth factors, cytokines and extracellular vehicles (EVs). EVs are multi-signal messengers that play a serious role in intercellular signaling through carrying cargo such as mRNA, miRNA, and proteins. Numerous studies have shown that MSC-derived EVs are able to mimic the effects of the cell of origin on immune cells. In this review, we discuss the current studies dealing with MSC-based therapies in autoimmune diseases and provide a vision and highlight in order to introduce MSC-derived EVs as an alternative and emerging modality for autoimmune disorders.

Dietary salt promotes ischemic brain injury and is associated with parenchymal migrasome formation

Sodium chloride promotes vascular fibrosis, arterial hypertension, pro-inflammatory immune cell polarization and endothelial dysfunction, all of which might influence outcomes following stroke. But despite enormous translational relevance, the functional importance of sodium chloride in the pathophysiology of acute ischemic stroke is still unclear. In the current study, we show that high-salt diet leads to significantly worse functional outcomes, increased infarct volumes, and a loss of astrocytes and cortical neurons in acute ischemic stroke. While analyzing the underlying pathologic processes, we identified the migrasome as a novel, sodium chloride-driven pathomechanism in acute ischemic stroke. The migrasome was previously described in vitro as a migrating organelle, which incorporates and dispatches cytosol of surrounding cells and plays a role in intercellular signaling, whereas a pathophysiological meaning has not been elaborated. We here confirm previously reported characteristics of the migrasome in vivo. Immunohistochemistry, electron microscopy and proteomic analyses further demonstrate that the migrasome incorporates and dispatches cytosol of surrounding neurons following stroke. The clinical relevance of these findings is emphasized by neuropathological examinations, which detected migrasome formation in infarcted brain parenchyma of human stroke patients. In summary, we demonstrate that high-salt diet aggravates stroke outcomes, and we characterize the migrasome as a novel mechanism in acute stroke pathophysiology.

Fluorimetric Studies of a Transmembrane Protein and Its Interactions with Differently Functionalized Silver Nanoparticles

Transmembrane proteins play important roles in intercellular signaling to regulate interactions among the adjacent cells and influence cell fate. The study of interactions between membrane proteins and nanomaterials is paramount for the design of nanomaterial-based therapies. In the present work, the fluorescence properties of the transmembrane receptor Notch2 have been investigated. In particular, the steady-state and time-resolved fluorescence methods have been used to characterize the emission of tryptophan residues of Notch2 and then this emission is used to monitor the effect of silver colloids on protein behavior. To this aim, silver colloids are prepared with two different methods to make sure that they bear hydrophilic (citrate ions, C-AgNPs) or hydrophobic (dodecanethiol molecules, D-AgNPs) capping agents. The preparation procedures are tightly controlled to obtain metal cores with similar size distributions (7.4 ± 2.5 and 5.0 ± 0.8 nm, respectively), thus, making the comparison of the results easier. The occurrence of strong interactions between Notch2 and D-AgNPs is suggested by the efficient and statistically relevant quenching of the stationary protein emission already at low nanoparticle (NP) concentrations (ca. 12% quenching with [D-AgNPs] = 0.6 nM). The quenching becomes even more pronounced (ca. 60%) when [D-AgNPs] is raised to 8.72 nM. On the other hand, the addition of increasing concentrations of C-AgNPs to Notch2 does not affect the protein fluorescence (intensity variations below 5%) indicating that negligible interactions are taking place. The fluorescence data, recorded in the presence of increasing concentrations of silver nanoparticles, are then analyzed through the Stern-Volmer equation and the sphere of action model to discuss the nature of interactions. The effect of D-AgNPs on the fluorescence decay times of Notch2 is also investigated and a decrease in the average decay time is observed (from 4.64 to 3.42 ns). The observed variations of the stationary and time-resolved fluorescence behavior of the protein are discussed in terms of static and collisional interactions. These results document that the capping shell is able to drive the protein-particle interactions, which likely have a hydrophobic nature.

Exosomes in Acquired Neurological Disorders: New Insights into Pathophysiology and Treatment.

Exosomes are endogenous nanovesicles that play critical roles in intercellular signaling by conveying functional genetic information and proteins between cells. Exosomes readily cross the blood-brain barrier and have promise as therapeutic delivery vehicles that have the potential to specifically deliver molecules to the central nervous system (CNS). This unique feature also makes exosomes attractive as biomarkers in diagnostics, prognostics, and therapeutics in the context of multiple significant public health conditions, including acquired neurological disorders. The purpose of this review is to summarize the state of the science surrounding the relevance of extracellular vesicles (EVs), particularly exosomes, to acquire neurological disorders, specifically traumatic brain injury (TBI), spinal cord injury (SCI), and ischemic stroke. In total, ten research articles were identified that examined exosomes in the context of TBI, SCI, or stroke; these manuscripts were reviewed and synthesized to further understand the current role of exosomes in the context of acquired neurological disorders. Of the ten published studies, four focused exclusively on TBI, one on both TBI and SCI, and five on ischemic stroke; notably, eight of the ten studies were limited to pre-clinical samples. The present review is the first to discuss the current body of knowledge surrounding the role of exosomes in the pathophysiology, diagnosis, and prognosis, as well as promising therapeutic strategies in TBI, SCI, and stroke research.

Exosomes: A Paradigm in Drug Development against Cancer and Infectious Diseases

Extracellular vesicles are small single lipid membrane entity secreted by eukaryotic and prokaryotic cells and play an important role in intercellular signaling and nutrient transport. The last few decades have witnessed a plethora of research on these vesicles owing to their ability to answer many hidden facts at the supramolecular level. These extracellular vesicles have attracted the researchers because they act as shuttle agents to transfer biomolecules/drugs between cells. Recently, studies have shown the application of exosomes in tumor therapy and infectious disease control. The present review article shows the importance of exosomes in cancer biology and infectious disease diagnoses and therapy and provides comprehensive account of exosomes biogenesis, extraction, molecular profiling, and application in drug delivery.

* Extracellular Vesicles and Their Versatile Roles in Tissue Engineering.

The importance of extracellular vesicles (EVs) - especially exosomes - in tissue engineering continues to increase due to their pivotal roles in intercellular signaling in a plethora of biological ...

TP53 Mutational Status and ROS Effect the Expression of the Survivin-Associated Radio-Adaptive Response.

A survivin-associated radio-adaptive response, characterized by increased radiation resistance or sensitization, was induced by exposure to 5 mGy of ionizing radiation and was correlated to the TP53 mutational status of exposed cells. Ten human cancer lines were investigated: colorectal carcinomas HCT116 and RKO [TP53 wild-type (WT)] and their respective TP53 null isogenic lines; breast adenocarcinomas MCF7 (TP53 WT) and MDA-MB-231 (TP53 Mut); lung carcinomas A549 (TP53 WT) and NCI-H1975 (TP53 Mut); and pancreatic carcinomas Hs766T (TP53 WT) and Panc-1 (TP53 Mut). Radiation induced (5 mGy) changes in the subsequent responses to 2 Gy in a multi-dose paradigm. Effects on radiation sensitivity were associated with changes in survivin's intracellular translocation to the cytoplasm (TP53 WT) or nucleus (TP53 Mut). Survival responses were determined using a colony forming assay. Intracellular localization of survivin was determined by ELISA and correlated with survival response. Two 2 Gy doses had minimal effects on the intracellular translocation of survivin. When preceded 15 min earlier by a 5 mGy exposure, survivin translocated to the cytoplasm in all of the TP53 WT cell lines, and to the nuclei in the TP53 null and Mut cells. All TP53 WT cells were protected (P < 0.001) by 5 mGy exposures, while Mut cells were sensitized (P < 0.001). HCT116 and RKO TP53 WT cells were admixed with their respective isogenic TP53 null counterparts in different proportions: 75% to 25%, 50% to 50% and 25% to 75%, respectively. All mixed confluent cultures expressed enhanced radio-sensitization (P ≤ 0.047) characteristic of TP53 Mut cells, which could be inhibited by their exposure to the antioxidant N-acetyl-l-cysteine (NAC) indicating a role for intercellular signaling by reactive oxygen species (ROS). ROS signaling in propagating the survivin-mediated response is involved in both intra- and intercellular communication processes.

Exosomes in Toxicology: Relevance to Chemical Exposure and Pathogenesis of Environmentally Linked Diseases.

Chronic exposure to environmental toxins has been known to initiate or aggravate various neurological disorders, carcinomas and other adverse health effects. Uptake by naïve cells of pathogenic factors such as danger-associated molecules, mRNAs, miRNAs or aggregated proteins leads to disruption in cellular homeostasis further resulting in inflammation and disease propagation. Although early research tended to focus solely on exosomal removal of unwanted cellular contents, more recent reports indicate that these nano-vesicles play an active role in intercellular signaling. Not only is the exosomal cargo cell type-specific, but it also differs between healthy and dying cells. Moreover, following exosome uptake by naïve cells, the contents from these vesicles can alter the fate of recipient cells. Since exosomes can traverse long distances, they can influence distantly located cells and tissues. This review briefly explores the role played by environmental toxins in stimulating exosome release in the context of progressive neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's, as well as certain cancers such as lung, liver, ovarian, and tracheal carcinomas.

The clinical significance of platelet microparticle-associated microRNAs.

Circulating blood platelets play a central role in the maintenance of hemostasis. They adhere to subendothelial extracellular matrix proteins that become exposed upon vessel wall damage, which is followed by platelet activation, further platelet recruitment, platelet aggregation and formation of an occlusive, or non-occlusive, platelet thrombus. Platelets host a surprisingly diverse transcriptome, which is comprised of ~9500 messenger RNAs (mRNAs) and different classes of non-coding RNAs, including microRNAs, as well as a significant repertoire of proteins that contribute to their primary (adhesion, aggregation, granule secretion) and alternative (RNA transfer, mRNA translation, immune regulation) functions. Platelets have the propensity to release microparticles (MPs; 0.1-1 μm in diameter) upon activation, which may mediate inflammatory responses and contribute to exacerbate inflammatory diseases and conditions. Carrying components of the platelets' cytoplasm, platelet MPs may exert their effects on recipient cells by transferring their content in platelet-derived bioactive lipid mediators, cytokines, mRNAs and microRNAs. Platelet MP-associated microRNAs may thus function also outside of platelets and play an important role in intercellular signaling and gene expression programming across the entire circulatory system. The role and importance of platelet MP-associated microRNAs in various aspects of biology and pathophysiology are increasingly recognized, and now provide the scientific basis and rationale to support further translational research and clinical studies. The clinical significance, pathophysiological role as well as the diagnostic and therapeutic potential of platelet MP-associated microRNAs in cardiovascular diseases, platelet transfusion and cancer will be discussed.

Abstract A05: Proteomic analysis of exosome content changes induced by gemcitabine in pancreatic cancer stromal cells

Abstract Despite recent advances in multimodality therapy, pancreatic ductal adenocarcinoma (PDAC) remains the 3rd leading cause of cancer-related deaths in the United States with a 5-year survival rate of 8%. One hallmark of this disease is desmoplasia that consist largely of stromal cells and extracellular matrix (ECM) proteins. The abundant stromal cells provide a unique tumor microenvironment that aid tumorigenesis. Unfortunately, there is little data on the mechanism underlying the stromal cells’ ability to impact chemoresistance in neighboring cancer cells in response to drug treatment. Exosomes are small vesicles secreted into the extracellular environment from cells and evidence is accumulating that exosomes play a key role in intercellular signaling. Researchers have focused on exosomes secreted from cancer cells, but the function of stromal exosomes has not been investigated, especially in pancreatic cancer. As stromal cells comprise the majority of the pancreatic tumor bulk, we hypothesize that stromal cell-derived exosomes help pancreatic cancer cells resist chemotherapy. Thus, it is critical to characterize the complex composition of stromal exosomes. In this study, we analyzed the proteomic content of exosomes collected from stromal cell lines created from PDAC patients. Furthermore, we also determined the change in exosomal proteomic composition after the patient-derived stromal cells were treated with gemcitabine, the current standard of care chemotherapy for patients with PDAC. Our studies have identified possible diagnostic and prognostic biomarkers and suggest novel molecular mechanism responsible for the tumor microenvironment’s ability to regulate chemoresistance in neighboring pancreatic cancer cells. Citation Format: Wei Huang, Peter Feist, Amanda Hummon, Reginald Hill.{Authors}. Proteomic analysis of exosome content changes induced by gemcitabine in pancreatic cancer stromal cells. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr A05.

Extracellular Vesicles Isolated from the Leaf Apoplast Carry Stress-Response Proteins.

Exosomes are extracellular vesicles (EVs) that play a central role in intercellular signaling in mammals by transporting proteins and small RNAs. Plants are also known to produce EVs, particularly in response to pathogen infection. The contents of plant EVs have not been analyzed, however, and their function is unknown. Here, we describe a method for purifying EVs from the apoplastic fluids of Arabidopsis (Arabidopsis thaliana) leaves. Proteomic analyses of these EVs revealed that they are highly enriched in proteins involved in biotic and abiotic stress responses. Consistent with this finding, EV secretion was enhanced in plants infected with Pseudomonas syringae and in response to treatment with salicylic acid. These findings suggest that EVs may represent an important component of plant immune responses.

Abstract 4971: Detection of receptor and non receptor tyrosine kinases in the culture filtrates of astrocytes and cancer cells

Abstract As demonstrated in our previous work, meningioma cells shed their membrane receptors and these receptors function in a cell-free environment. The purpose of this study was to determine if receptor and non-receptor kinases were similarly being released in different cancers. Using antibody arrays, cell culture filtrates from glioma and prostate cancer cell lines were assessed to determine if solubilized receptors and kinases were present. Supernatants from cells treated with Brefeldin-A were analyzed similarly. Further, culture filtrates concentrated by ultrafiltration were subjected to luminescence based kinase assays and western blotting to assess the active forms. Cell viability was measured with Trypan blue staining in all the experiments. Our results demonstrate that an array of receptors are solubilized into culture filtrates of brain and prostate cancer cell lines within the first 24 hours. Simultaneously, soluble receptors in the culture filtrates of astrocytes within 48 hours of culture were also detected. Compared to astrocytes, cancer cells showed elevated levels of receptors and kinases released at 48 hours. As many as 15 receptor or non-receptor tyrosine kinases including Src family of kinases were found in the culture filtrates of the brain and prostate cancer cell lines. Treatment with Brefeldin-A inhibited the release of receptors and kinases partially, but markedly, within first 24 hours suggesting a secretory pathway for their release. Additionally, phosphorylation of tyrosine kinases was detected in the western blots within 24 hours. Kinase assays further confirmed active nature of the extracellular kinases, particularly, when checked for the active form of Lyn kinase in the culture filtrates, phosphorylation on Tyrosine-507 was detected along with the total forms. Cell viability assays on the cultured cells revealed more than 98% viability, thus eliminating the possibility of the release of detected kinases from dead cells. In light of this study, we propose a new niche for solubilized receptor and tyrosine kinase functioning that might have a role in intercellular signaling in cell-free conditions both in disease and physiological statuses. Note: This abstract was not presented at the meeting. Citation Format: Venkateswara Rao Gogineni. Detection of receptor and non receptor tyrosine kinases in the culture filtrates of astrocytes and cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4971. doi:10.1158/1538-7445.AM2015-4971

Exosome-like vesicles derived by Schistosoma japonicum adult worms mediates M1 type immune- activity of macrophage.

Exosomes are 30-100-nm membrane vesicles of endocytic origin that are released into the extracellular space upon fusion of the multi-vesicular bodies (MVB) with the plasma membrane, while initial studies described that the role of exosomes was a reticulocyte cargo-disposal mechanism allowing remodeling of the plasma membrane during the maturation of reticulocytes to erythrocytes. Recent studies indicate that exosomes are secreted by most cells and pathogens and play an important role in intercellular signaling and exert regulatory function by carrying bioactive molecules. As numerous pathogens, adult worm of Schistosoma japonicum (S. japonicum) reside in mesenteric veins of definitive host including man and mammal animals. It was reported that the worms or the eggs also have specialized secretion systems to export effector proteins or other molecules into host target cells. However, the mechanisms involved remained unclear. This study investigated the isolation of the exosome-like vesicles secreted by S. japonicum adult worms and its immune activity on microphage in vitro. In this report, we identified exosome-based secretion as a new mechanism for protein secretion by S. japonicum. Electron microscopy tomography revealed the previously unidentified ultrastructural detail of exosome-like vesicles with high resolution; they were found to be typical spherical shape and to have a diverse population that varies in size of 30-100nm. Exosome-like vesicles isolated from S. japonicum contained a significantly different protein compared with debris pelleted and the apoptosis body. We also demonstrate that macrophages were preferentially differentiated into the M1 subtype while being treated with S. japonicum exosome-like vesicles. This study reveals there are exosome-like vesicles derived by S. japonicum adult worms, and the exosome-like vesicles can mediate M1-type immune- activity of macrophage.

Exosomes secreted from GATA-4 overexpressing mesenchymal stem cells serve as a reservoir of anti-apoptotic microRNAs for cardioprotection

BackgroundExosomes play an important role in intercellular signaling and exert regulatory function by carrying bioactive molecules. This study investigated (1) the cardioprotective capabilities of exosomes derived from mesenchymal stem cells (MSCs) overexpressing GATA-4 (MSCGATA-4) and (2) its underlying regulatory mechanisms for expression of target proteins in recipient cells. Methods and resultsExosomes were isolated and purified from MSCGATA-4 (ExoGATA-4) and control MSCs (ExoNull). Cell injury was investigated in primary cultured rat neonatal cardiomyocytes (CM) and in the rat heart. Exosomes contributed to increased CM survival, reduced CM apoptosis, and preserved mitochondrial membrane potential in CM cultured under a hypoxic environment. Direct intramyocardial transplantation of exosomes at the border of an ischemic region following ligation of the left anterior descending coronary artery significantly restored cardiac contractile function and reduced infarct size. Real-time PCR revealed that several anti-apoptotic miRs were highly expressed in ExoGATA-4. Rapid internalization of ExoGATA-4 by CM was documented using time-lapse imaging. Subsequent expression of these miRs, particularly miR-19a was higher in CM and in the myocardium treated with ExoGATA-4 compared to those treated with ExoNull. The enhanced protective effects observed in CM were diminished by the inhibition of miR-19a. The expression level of PTEN, a predicted target of miR-19a, was reduced in CM treated with ExoGATA-4, which resulted in the activation of the Akt and ERK signaling pathways. ConclusionsExoGATA-4 upon transplantation in the damaged tissue mediate protection by releasing multiple miRs responsible for activation of the cell survival signaling pathway.

Migrasomes: a new organelle of migrating cells.

Cell migration is a multi-step process that involves the coordinated action of signaling networks, cytoskeletal dynamics and vesicular trafficking, leading to protrusion and adhesion at the leading edge of cells and contraction and detachment at their rear. In a recent paper in Cell Research, Ma et al. describe the biogenesis of a new exosome-like organelle--named migrasomes--that derive from retraction fibers at the rear of migrating cells and their potential roles in inter-cellular signaling.

Isolated human uterine telocytes: immunocytochemistry and electrophysiology of T-type calcium channels.

Recently, telocytes (TCs) were described as a new cell type in the interstitial space of many organs, including myometrium. TCs are cells with very long, distinctive extensions named telopodes (Tps). It is suggested that TCs play a major role in intercellular signaling, as well as in morphogenesis, especially in morphogenetic bioelectrical signaling. However, TC plasma membrane is yet unexplored regarding the presence and activity of ion channels and pumps. Here, we used a combination of in vitro immunofluorescence and patch-clamp technique to characterize T-type calcium channels in TCs. Myometrial TCs were identified in cell culture (non-pregnant and pregnant myometrium) as cells having very long Tps and which were positive for CD34 and platelet-derived growth factor receptor-α. Immunofluorescence analysis of the subfamily of T-type (transient) calcium channels CaV3.1 and CaV3.2 presence revealed the expression of these ion channels on the cell body and Tps of non-pregnant and pregnant myometrium TCs. The expression in TCs from the non-pregnant myometrium is less intense, being confined to the cell body for CaV3.2, while CaV3.1 was expressed both on the cell body and in Tps. Moreover, the presence of T-type calcium channels in TCs from non-pregnant myometrium is also confirmed by applying brief ramp depolarization protocols. In conclusion, our results show that T-type calcium channels are present in TCs from human myometrium and could participate in the generation of endogenous bioelectric signals responsible for the regulation of the surrounding cell behavior, during pregnancy and labor.

Experimental Investigations of Single Liposome to Supported Bilayer Binding Events

One of the primary ways in which cells interact with their environments is by release of extracellular vesicles. These cell-derived submicron sized containers are formed either from the cell plasma membrane (microvesicles) or secreted from multivesicular bodies (exosomes). These vesicles contain nucleic acids and proteins that have been suggested to play an important role in intercellular signaling and the process of molecular communication between cells. In microvesicle-mediated intercellular communication, vesicles released by a donor cell must bind to the plasma membrane of a recipient cell in order to deliver their cargo to the target. Despite the important physiological role of vesicle-plasma membrane fusion and vesicle endocytosis, the details of the physical interactions between microvesicles and the plasma membrane are still poorly understood. To better understand the forces which occur between microvesicles and cells, we experimentally investigate single liposome binding events with a supported lipid bilayer. We employed total internal reflection fluorescence (TIRF) microscopy to observe liposome interactions with a supported lipid bilayer, and compare the experimentally observed bond lifetimes to the theoretical estimates. Our experimental observations indicate that bond lifetimes increase with both increasing liposome size and decreasing temperature.

Extracellular vesicle-mediated transfer of long non-coding RNA ROR modulates chemosensitivity in human hepatocellular cancer

Hepatocellular cancers (HCC) are highly resistant to chemotherapy. TGFβ has been associated with chemoresistance in some human cancers but the mechanisms involved are unknown. We explored how TGFβ might contribute to altered responses to therapy by assessing the involvement and mechanistic contribution of extracellular vesicle long non-coding RNA (lncRNA) in mediating TGFβ-dependent chemoresistance. TGFβ reduced the sensitivity of HCC cells to sorafenib or doxorubicin and altered the release of both extracellular vesicles and of selected lncRNA within these vesicles. Amongst these, lincRNA-ROR (linc-ROR), a stress-responsive lncRNA was highly expressed in HCC cells and enriched within extracellular vesicles derived from tumor cells. Incubation with HCC-derived extracellular vesicles increased linc-ROR expression and reduced chemotherapy-induced cell death in recipient cells. Sorafenib increased linc-ROR expression in both tumor cells and extracellular vesicles, whereas siRNA to linc-ROR increased chemotherapy-induced apoptosis and cytotoxicity. Tumor-initiating cells that express CD133 have an increased resistance to therapy. TGFβ increased expression of CD133+ cells and colony growth in limiting dilution assays, both of which were attenuated by linc-ROR knockdown. These data provide mechanistic insights into primary chemoresistance in HCC by showing that: (a) TGFβ selectively enriches linc-RoR within extracellular vesicles, which has a potential role in intercellular signaling in response to TGFβ; (b) expression and enrichment of linc-ROR during chemotherapeutic stress plays a functional role in chemoresistance; and (c) the effects of TGFβ on chemoresistance in HCC may involve linc-RoR-dependent effects on tumor-initiating cells. These findings implicate extracellular vesicle lncRNA as mediators of the chemotherapeutic response, and support targeting linc-ROR to enhance chemosensitivity in HCC.

ABC Transporters in Dictyostelium discoideum Development

ATP-binding cassette (ABC) transporters can translocate a broad spectrum of molecules across the cell membrane including physiological cargo and toxins. ABC transporters are known for the role they play in resistance towards anticancer agents in chemotherapy of cancer patients. There are 68 ABC transporters annotated in the genome of the social amoeba Dictyostelium discoideum. We have characterized more than half of these ABC transporters through a systematic study of mutations in their genes. We have analyzed morphological and transcriptional phenotypes for these mutants during growth and development and found that most of the mutants exhibited rather subtle phenotypes. A few of the genes may share physiological functions, as reflected in their transcriptional phenotypes. Since most of the abc-transporter mutants showed subtle morphological phenotypes, we utilized these transcriptional phenotypes to identify genes that are important for development by looking for transcripts whose abundance was unperturbed in most of the mutants. We found a set of 668 genes that includes many validated D. discoideum developmental genes. We have also found that abcG6 and abcG18 may have potential roles in intercellular signaling during terminal differentiation of spores and stalks.

A gene encoding a peptide with similarity to the plant IDA signaling peptide (AtIDA) is expressed most abundantly in the root-knot nematode (Meloidogyne incognita) soon after root infection

Small peptides play important roles in intercellular signaling. Inflorescence deficient in abscission (ida) is an Arabidopsis mutant that does not abscise (shed) its flower petals. The IDA gene encodes a small, secreted peptide that putatively binds to two redundant receptor-like kinases (HAESA and HAESA-like2) that initiate a signal transduction pathway. We identified IDA-like (IDL) genes in the genomic sequence for Meloidogyne incognita and Meloidogyne hapla. No orthologous sequences were found in any other genus of nematodes. Transcript for both M. incognita and M. hapla IDLs were found in total RNA isolated from infected root systems of tomato, Solanum lycopersicum. Five and three prime RACE of RNA from M. incognita infected tomato roots revealed a sequence of 392 nt that includes a poly (A) tail of 39 nt. The open reading frame encodes a 47 aa protein with a putative 25 aa N-terminal signal peptide. Expression of MiIDL1 is very low in eggs and pre-parasitic J2 and rapidly increases in the first four days post inoculation (dpi) and then declines at approximately 14 dpi. A proposed role for the root-knot nematode IDL is discussed.