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PubMed1986

Alpha 1-adrenergic stimulation and cytoplasmic free calcium concentration in cultured renal proximal tubular cells: evidence for compartmentalization of quin-2 and fura-2.

This study was designed to examine the role of changes in cytoplasmic free calcium concentration ([Ca2+]i) during the response to alpha 1-adrenergic agonists in cultured renal proximal tubular cells. Experiments were carried out on primary cultures of canine proximal tubular cells grown in defined culture medium on a solid support, on collagen-coated polycarbonate membranes, or on collagen-coated glass coverslips. Quin-2 and fura-2 were used to monitor [Ca2+]i. The basal level of [Ca2+]i was 101 nM, as measured with quin-2, and 122 nM, as determined using fura-2. Fluorescence flow cytometry revealed that about 85% of the population of proximal tubular cells responded to phenylephrine with an increase in [Ca2+]i. Phenylephrine (10(-5) M) caused an immediate actual increase in [Ca2+]i by 18 and 24%, as determined with quin-2 and fura-2, respectively, with the peak increase in [Ca2+]i averaging 22% and 44% over the basal level (180-300 sec). This effect did not require extracellular calcium. The effect of phenylephrine was abolished by prazosin and verapamil. Fluorescence microscopy of quin-2 or fura-2 loaded cells revealed punctate areas of fluorescence within the cytoplasm suggesting vesicular uptake of the dyes. Pinocytotic entrapment of the dyes was demonstrated by the transfer of cell-impermeant fura-2 across tubular cell monolayers mounted in Ussing chambers. The transfer of the dye was similar to that of a marker of fluid-phase pinocytosis, Lucifer Yellow (LY). This pinocytotic entrapment of Ca2+-indicators would lead to underestimation of the actual calcium transients. Microfluorometric study of single proximal tubular cells "scrape-loaded" with fura-2 revealed a four-fold increase in [Ca2+]i concentration following stimulation with phenylephrine.

Aminoquinolines↗
PubMed2005

Evidence in support of a role for plant-mediated proteolysis in the rumens of grazing animals.

The present work aimed to differentiate between proteolytic activities of plants and micro-organisms during the incubation of grass in cattle rumens. Freshly cut ryegrass was placed in bags of varying permeability and incubated for 16 h in the rumens of dairy cows that had previously grazed a ryegrass sward, supplemented with 4 kg dairy concentrate daily. Woven polyester bags (50 microm pore size) permitted direct access of the micro-organisms and rumen fluid enzymes to the plant material. The polythene was impermeable even to small molecules such as NH(3). Dialysis tubing excluded micro-organisms and rumen enzymes/metabolites larger than 10 kDa. DM loss was 46.3 % in polyester, 36.2 % in polythene and 38.1 % in dialysis treatments. It is possible that the DM loss within polythene bags occurred due to a solubilisation of plant constituents (e.g. water-soluble carbohydrates) rather than microbial attachment/degradation processes. The final protein content of the herbage residues was not significantly different between treatments. Regardless of bag permeability, over 97 % of the initial protein content was lost during incubations in situ. Electrophoretic separation showed that Rubisco was extensively degraded in herbage residues whereas the membrane-associated, light-harvesting protein remained relatively undegraded. Protease activity was detected in herbage residues and bathing liquids after all incubation in situ treatments. Although rumen fluid contains proteases (possibly of plant and microbial origin), our results suggest that, owing to cell compartmentation, their activity against the proteins of intact plant cells is limited, supporting the view that plant proteases are involved in the degradation of proteins in freshly ingested herbage.

Animal Nutritional Physiological Phenomena↗
PubMed1997

Compartmentalized vesicular traffic around the hair cell cuticular plate.

Through thin-section and freeze-fracture electron microscopy, we identify structural correlates of an intense vesicular traffic in a narrow band of cytoplasm around the cuticular plate of the bullfrog vestibular hair cells. Myriads of coated and uncoated vesicles associated with longitudinally oriented microtubules populate the narrow cytoplasmic region between the cuticular plate and the actin network of the apical junctional belt. If microtubules in the sensory hair cells, like those in axons, are pathways for organelle transport, then the characteristic distribution of microtubules around the cuticular plate represents transport pathways across the apical region of the hair cells. This compartmentalized membrane traffic system appears to support an intense vesicular release and uptake along a band of apical plasma membrane near the cell border. Functions of this transport system may include membrane recycling as well as exocytotic and endocytotic exchange between the hair cell cytoplasm and the endolymphatic compartment.

Acoustic Maculae↗
PubMed1996

Analysis of the role of prespore gene expression in the compartmentalization of mother cell-specific gene expression during sporulation of Bacillus subtilis.

A hallmark of sporulation of Bacillus subtilis is the formation of two distinct cells by an asymmetric division. The development programs in these two cells involve the compartmentalized activities of sigma E in the larger mother cell and of sigma F in the smaller prespore. Activation of sigma E requires expression of the sigma F-directed gene spoIIR. By immunofluorescence microscopy of a strain containing a spoIIR-lacZ fusion, we have shown that spoIIR is transcribed exclusively in the prespore. By placing spoIIR under the control of PspoIIE, it was possible to express spoIIR before the spore septum was formed. Strains containing the PspoIIE-spoIIR construct activated sigma E only in the mother cell in organisms that underwent the asymmetric sporulation division. Thus, compartmentalization of sigma E activity did not require the compartmentalization of spoIIR expression. Nor did the compartmentalization of sigma E require SpoIIAA, SpoIIAB, sigma F, or sigma F-dependent transcription, all of which are required for prespore-specific gene expression. It is inferred that although sigma F and sigma E direct compartmentalized gene expression, neither of these sigma factors, nor the genes under their control, directs the process of compartmentalization.

Bacillus subtilis↗
PubMed1987

Recognition of specific targets by cultured dorsal root ganglion neurons.

We have assessed the effects of different target cell populations on axonally transported proteins by the use of compartmental cell culture systems that separate the soma from the growing axons of rat sensory neurons. The labeling of 3 rapidly transported proteins diminishes when the growing axon contacts spinal cord cells (which are normal in vivo targets), and remains unaffected by contact with fibroblasts or heart cells. Medium conditioned by spinal cord cells does not exert this effect. Thus, specific classes of cells may be distinguished as target tissue by sensory neurons in vitro. Such recognition is accompanied by specific molecular changes in axonally transported proteins.

Animals↗
PubMed1988

Phospholipase A2 activity in resting and activated human neutrophils. Substrate specificity, pH dependence, and subcellular localization.

We have studied the phospholipase A2 activity in fractionated human neutrophils, employing labeled phosphatidylinositol, phosphatidylcholine, and phosphatidylethanolamine as exogenous substrates. We used these phospholipid substrates labeled in the sn-1 position and measured the resulting labeled lysophospholipid forms in order to ascertain the phospholipase A2 specificity. In postnuclear supernatants from resting and A23187-activated cells, the phospholipase A2 activity showed a similar pH dependence curve with two pH optima at 5.5 and 7.5. Extracts from activated cells showed a 3-6-fold increase in enzyme activity. The subcellular distribution of phospholipase A2 activity in resting and A23187-treated human neutrophils was investigated by fractionation of postnuclear supernatants on continuous sucrose gradients. The neutral phospholipase A2 behaved as a membrane-bound enzyme and was mainly localized in the plasma membrane, the azurophilic granule, and in an ill-defined region of the gradient between the specific granules and mitochondria. The phospholipase A2 located in this undefined region showed a higher degree of activation than that located in other subcellular particulates in A23187-treated cells. This specific activation of an intracellular phospholipase A2 activity during cell stimulation indicates that cell compartmentalization may play a role in the formation of cell-activating and/or signal-transducing agents through the generation of arachidonate metabolites. Phosphatidylinositol was a better substrate for the plasma membrane enzyme, whereas phosphatidylcholine and phosphatidylethanolamine behaved as better substrates for intracellular organelle phospholipase A2 activities. The phospholipase A2 with maximal activity at pH 5.5 behaved as a soluble enzyme, and was almost completely localized in the azurophilic granules. Upon cell activation this acid enzyme activity was released in a similar way to beta-glucuronidase, a marker of azurophilic granules. These results demonstrate the different molecular properties of the phospholipase A2 activity, on the basis of its cellular location.

Calcimycin↗
PubMed1984

The mononuclear cells of human mesenteric blood, intestinal mucosa and mesenteric lymph nodes: compartmentalization of NK cells.

The proportions of T cell subsets and Leu 7+ cells and the spontaneous cell-mediated cytotoxicity (SCMC) of isolated mononuclear cells have been determined across the mesenteric vascular bed and along the intestinal mucosal-mesenteric lymph node (MLN) axis in patients undergoing abdominal surgery. Whereas the proportion of T4+ and T8+ cells were similar in simultaneously taken PVB and mesenteric venous blood (MVB), the proportion of Leu 7+ cells was higher in MVB in 16 of 17 studies (15.4 +/- 6.8%, 10.8 +/- 5.1%). Additional studies showed that the proportions of lymphocyte subsets in peripheral arterial blood are the same as those in PVB. Thus, an enrichment of Leu 7+ cells occurs across the mesenteric vascular bed. Isolated intestinal and MLN mononuclear cells contained similarly high proportions of T4+ and T8+ cells as in PVB but Leu 7+ cells made up a minority subpopulation in intestinal (1.3 +/- 0.8%) and MLN mononuclear cells (1.0 +/- 0.9%). The SCMC of intestinal and MLN mononuclear cells was low and paralleled the proportion of Leu 7+ cells. Despite the higher proportions of Leu 7+ cells in MVB, the SCMC was less than that of PVB in eight patients with inflamed intestine and not significantly different from PVB in seven patients with normal intestines. These paradoxical findings were at least in part due to inhibitory factors in mesenteric plasma. In conclusion, NK cells appear to be largely confined within the vascular system and the enrichment of Leu 7+ cells across the mesenteric vascular bed suggests that this compartmentalization may be due to differences in the traffic of lymphocyte subpopulations through the intestinal mucosa and MLN.

Cell Movement↗
PubMed2004

Computerized microfluidic cell culture using elastomeric channels and Braille displays.

Computer-controlled microfluidics would advance many types of cellular assays and microscale tissue engineering studies wherever spatiotemporal changes in fluidics need to be defined. However, this goal has been elusive because of the limited availability of integrated, programmable pumps and valves. This paper demonstrates how a refreshable Braille display, with its grid of 320 vertically moving pins, can power integrated pumps and valves through localized deformations of channel networks within elastic silicone rubber. The resulting computerized fluidic control is able to switch among: (i) rapid and efficient mixing between streams, (ii) multiple laminar flows with minimal mixing between streams, and (iii) segmented plug-flow of immiscible fluids within the same channel architecture. The same control method is used to precisely seed cells, compartmentalize them into distinct subpopulations through channel reconfiguration, and culture each cell subpopulation for up to 3 weeks under perfusion. These reliable microscale cell cultures showed gradients of cellular behavior from C2C12 myoblasts along channel lengths, as well as differences in cell density of undifferentiated myoblasts and differentiation patterns, both programmable through different flow rates of serum-containing media. This technology will allow future microscale tissue or cell studies to be more accessible, especially for high-throughput, complex, and long-term experiments. The microfluidic actuation method described is versatile and computer programmable, yet simple, well packaged, and portable enough for personal use.

Audiovisual Aids↗
PubMed2004

Expansion and contraction of the NK cell compartment in response to murine cytomegalovirus infection.

NK cells are capable of responding quickly to infectious challenge and contribute to the early defense against a wide variety of pathogens. Although the innate NK cell response to murine CMV (MCMV) has been extensively characterized, its resolution and the fate of the activated NK cell population remains unexplored. Herein, we characterize both the expansion and contraction phases of the NK cell response to MCMV. We demonstrate that NK cell recruitment into the immune response to MCMV infection is restricted to the first 3 days of infection and as the peripheral NK cell compartment expands, NK cells undergo accelerated phenotypic maturation. During the resolution of the immune response, NK cell compartmental contraction is marked by the selective death of responding NK cells. Additionally, throughout the infection, a naive NK cell pool that remains responsive to additional stimuli is actively maintained. These findings illustrate the plasticity of the NK cell compartment in response to pathogens and underscore the homeostatic maintenance of the resting peripheral NK cell pool.

Animals↗
PubMed2026

Mapping subcellular microenvironments using oligonucleotide-directed proximity labeling.

Understanding how cells compartmentalize their biomolecules into discrete structures remains one of the fundamental goals of cell biology. The proliferation of proximity labeling (PL) technologies has been invaluable toward this goal, enabling biochemical "dissection" of compartments that would be intractable to classical biochemical methods. While robust PL approaches have long been established for targeting proteins of interest, targeting nucleic acids-RNAs and genomic loci-has remained significantly more challenging. Here, we review recent advancements in the field that overcome this longstanding roadblock by using programmable DNA oligonucleotides to direct PL enzyme localization. These tools are already revealing new insights into the molecular architecture of cellular compartments that lie at the heart of gene expression. They also provide a foundation for developing a new generation of PL tools that exploit the modularity and programmability of oligonucleotide-based devices to enable precise spatiotemporal control at previously inaccessible targets and in challenging specimen types.

Oligonucleotides↗
PubMed1995

A novel leukointegrin, alpha d beta 2, binds preferentially to ICAM-3.

The leukocyte-restricted beta 2 (CD18) integrins mediate cell adhesion in a variety of events essential for normal immune function. Despite extensive research in this field, only three members of this integrin subfamily have been described: CD11a/CD18 (LFA-1), CD11b/CD18 (Mac-1), and CD11c/CD18 (p150,95). We have identified a cDNA encoding a fourth alpha chain, alpha d, that associates with CD18. The alpha d subunit is more closely related to CD11b and CD11c than to CD11a. This integrin is expressed at moderate levels on myelomonocytic cell lines and subsets of peripheral blood leukocytes, and more strongly on tissue-compartmentalized cells such as foam cells, specialized macrophages found in aortic fatty streaks that may develop into atherosclerotic lesions. The alpha d/CD18 molecule exhibits preferential recognition of ICAM-3 over ICAM-1.

Amino Acid Sequence↗
PubMed1991

Limited clonal heterogeneity of antigen-specific T cells localizing in the pleural space during mycobacterial infection.

To detect possible differences in phenotype and fine specificity for mycobacterial antigens between CD4-positive T cells from peripheral blood (PB) and from inflammatory sites, we identified four patients presenting with a mycobacterial pleural exudate (PE) rich in PPD-specific lymphocytes and with a negative skin test to tuberculin purified protein derivative (PPD) and a negative proliferative response of PB lymphocytes to PPD at the same time. Several weeks after chemotherapy, these patients converted to PPD responsiveness in the periphery, and PPD-specific clones could be generated from PB at this stage. The phenotypic comparison of PE lymphocytes and concomitant PB lymphocytes obtained before treatment showed an increase of CD8 cells and a high frequency of HLA-DR-positive activated T cells in PE. The frequency of tetanus toxoid-specific and Candida albicans-specific proliferating T cells was lower than that of PPD-specific cells in PE but not in PB. PPD-specific clones were derived initially from PE and from PB once the patients had converted to PPD responsiveness. The two sets of clones from each patient were compared for proliferative response to mycobacterial antigen clusters of defined molecular weight ranges. A large number of PE-derived clones (36%) responded to a fraction of 27 to 35 kDa, whereas only one clone from PB responded to the same fraction. The purified antigen P32 (32 kDa), a soluble mycobacterial protein, stimulated PE-derived clones that were responsive to the 37- to 27-kDa fraction but did not stimulate PB-derived clones. The data demonstrate that PE- and PB-derived lymphocytes differ both in phenotype and in fine specificity, suggesting a limited clonal heterogeneity of T cells localizing at the inflammatory site in tuberculous patients without a PPD response in the periphery. Therefore T cells compartmentalized at inflammatory sites provide information that is different from that provided by T cells in the periphery.

Antigens, Bacterial↗
PubMed2003

Molecular dissection of a yeast septin: distinct domains are required for septin interaction, localization, and function.

The septins are a family of cytoskeletal proteins present in animal and fungal cells. They were first identified for their essential role in cytokinesis, but more recently, they have been found to play an important role in many cellular processes, including bud site selection, chitin deposition, cell compartmentalization, and exocytosis. Septin proteins self-associate into filamentous structures that, in yeast cells, form a cortical ring at the mother bud neck. Members of the septin family share common structural domains: a GTPase domain in the central region of the protein, a stretch of basic residues at the amino terminus, and a predicted coiled-coil domain at the carboxy terminus. We have studied the role of each domain in the Saccharomyces cerevisiae septin Cdc11 and found that the three domains are responsible for distinct and sometimes overlapping functions. All three domains are important for proper localization and function in cytokinesis and morphogenesis. The basic region was found to bind the phosphoinositides phosphatidylinositol 4-phosphate and phosphatidylinositol 5-phosphate. The coiled-coil domain is important for interaction with Cdc3 and Bem4. The GTPase domain is involved in Cdc11-septin interaction and targeting to the mother bud neck. Surprisingly, GTP binding appears to be dispensable for Cdc11 function, localization, and lipid binding. Thus, we find that septins are multifunctional proteins with specific domains involved in distinct molecular interactions required for assembly, localization, and function within the cell.

Amino Acid Sequence↗
PubMed2004

Translocation of the inhibitor of apoptosis protein c-IAP1 from the nucleus to the Golgi in hematopoietic cells undergoing differentiation: a nuclear export signal-mediated event.

The caspase inhibitor and RING finger-containing protein cellular inhibitor of apoptosis protein 1 (c-IAP1) has been shown to be involved in both apoptosis inhibition and signaling by members of the tumor necrosis factor (TNF) receptor family. The protein is regulated transcriptionally (eg, is a target for nuclear factor-kappaB [NF-kappaB]) and can be inhibited by mitochondrial proteins released in the cytoplasm upon apoptotic stimuli. The present study indicates that an additional level of regulation of c-IAP1 may be cell compartmentalization. The protein is present in the nucleus of undifferentiated U937 and THP1 monocytic cell lines. When these cells undergo differentiation under phorbol ester exposure, c-IAP1 translocates to the cytoplasmic side of the Golgi apparatus. This redistribution involves a nuclear export signal (NES)-mediated, leptomycin B-sensitive mechanism. Using site-directed mutagenesis, we localized the functional NES motif in the caspase recruitment domain (CARD) of c-IAP1. A nucleocytoplasmic redistribution of the protein was also observed in human monocytes as well as in tumor cells from epithelial origin when undergoing differentiation. c-IAP1 does not translocate from the nucleus of cells whose differentiation is blocked (ie, in cell lines and monocytes from transgenic mice overexpressing B-cell lymphoma 2 [Bcl-2] and in monocytes from patients with chronic myelomonocytic leukemia). Altogether, these observations associate c-IAP1 cellular location with cell differentiation, which opens new perspectives on the functions of the protein.

Active Transport, Cell Nucleus↗
PubMed2006

Specific remodeling of splenic architecture by cytomegalovirus.

Efficient immune defenses are facilitated by the organized microarchitecture of lymphoid organs, and this organization is regulated by the compartmentalized expression of lymphoid tissue chemokines. Mouse cytomegalovirus (MCMV) infection induces significant remodeling of splenic microarchitecture, including loss of marginal zone macrophage populations and dissolution of T and B cell compartmentalization. MCMV preferentially infected the splenic stroma, targeting endothelial cells (EC) as revealed using MCMV-expressing green fluorescent protein. MCMV infection caused a specific, but transient transcriptional suppression of secondary lymphoid chemokine (CCL21). The loss of CCL21 was associated with the failure of T lymphocytes to locate within the T cell zone, although trafficking to the spleen was unaltered. Expression of CCL21 in lymphotoxin (LT)-alpha-deficient mice is dramatically reduced, however MCMV infection further reduced CCL21 levels, suggesting that viral modulation of CCL21 was independent of LTalpha signaling. Activation of LTbeta-receptor signaling with an agonistic antibody partially restored CCL21 mRNA expression and redirected transferred T cells to the splenic T cell zone in MCMV-infected mice. These results indicate that virus-induced alterations in lymphoid tissues can occur through an LT-independent modulation of chemokine transcription, and targeting of the LT cytokine system can counteract lymphoid tissue remodeling by MCMV.

Animals↗
PubMed1991

Relationship between the calcium-mobilizing action of inositol 1,4,5-trisphosphate in permeable AR4-2J cells and the estimated levels of inositol 1,4,5-trisphosphate in intact AR4-2J cells.

Various experimental strategies were employed in an effort to explain the previously reported [Horstman, Takemura & Putney (1988) J. Biol. Chem. 263, 15297-15303] paradoxically high levels of inositol 1,4,5-trisphosphate [(1,4,5)IP3] in resting and substance-P-stimulated AR4-2J cells. The concentration-effect curves for substance-P-induced [3H](1,4,5)IP3 formation in [3H]inositol-labelled cells and substance-P-induced increase in intracellular [Ca2+] were essentially superimposable, suggesting that formation of (1,4,5)IP3 is limiting for cellular Ca2+ mobilization. In electrically permeabilized AR4-2J cells, (1,4,5)IP3 and other inositol polyphosphates stimulated Ca2+ release with potencies similar to those reported for other cell types, including the parent pancreatic acinar cell. Compartmentalization of basal (1,4,5)IP3 was suggested by the fact that this material was stable in the presence of antimycin A, although this toxin completely blocked agonist stimulation of phospholipase C. However, subcellular fractionation as well as permeabilization of the cells with Staphylococcus aureus alpha-toxin failed to provide evidence for binding or sequestration of [3H](1,4,5)IP3 in AR4-2J cells. The density of (1,4,5)IP3 receptors in AR4-2J cells was not sufficiently large to impose non-linearity in the relationship between (1,4,5)IP3 concentration and (1,4,5)IP3-induced Ca2+ release. Thus the apparent high concentrations of (1,4,5)IP3 in resting and stimulated AR4-2J cells are not indicative of atypically low sensitivity or high concentration of (1,4,5)IP3 receptors, nor is there evidence for compartmentalization of (1,4,5)IP3 outside of the cytoplasm in these cells. It is possible that soluble factors in the cytoplasm of AR4-2J cells regulate the free concentration of (1,4,5)IP3 or the sensitivity of receptors to (1,4,5)IP3.

Animals↗
PubMed2001

Abnormal dispersion of a purkinje cell subset in the mouse mutant cerebellar deficient folia (cdf).

Purkinje cells of different molecular phenotypes subdivide the cortex of the cerebellum both rostrocaudally into parasagittal bands and mediolaterally into transverse zones. Superimposed on the Purkinje cell compartmentation, the cerebellar cortex is pleated into a reproducible array of lobes and lobules. During cerebellar development, Purkinje cell bands are formed through the rostrocaudal dispersal of embryonic clusters, triggered primarily by a Reelin-dependent signaling pathway. In the naturally occurring mouse mutant cerebellar deficient folia (cdf), there is a failure of Purkinje cell dispersion that results in widespread Purkinje cell ectopia in the adult. The ectopia is restricted primarily to that subset of Purkinje cells that does not express zebrin II/aldolase C and that forms ectopic clusters in among the cerebellar nuclei. Most Purkinje cells that express zebrin II are located normally in a monolayer. Thus, the cerebellum of cdf mutants has a failure of Purkinje cell dispersion that is confined primarily to a zebrin II-negative (zebrin II(-)) subpopulation. Despite the Purkinje cell ectopia, the parasagittal band organization of the cerebellum is still clear. The shortening of the cortex is distributed evenly over all lobules, with the result that transverse expression boundaries are relocated with respect to the lobules and fissures. The number of Purkinje cells in the cdf/cdf cerebellum is similar to the number in littermate controls. Therefore, it appears that the lesion in cdf results in the failure of a zebrin II(-) Purkinje cell subset to disperse either due to a cell intrinsic defect or due to an abnormal interaction between the Purkinje cells and either granule cells or afferent inputs.

Animals↗
PubMed1993

Morphological transformation, tumorigenicity and src-specific cytotoxic T-lymphocyte-mediated tumor immunity induced by murine 3T3 cells expressing src oncogenes encoding novel non-myristylated N-terminal domains.

We previously reported the development of a src-specific tumor regression system in chickens in which preinfection with rASV1702, a mutant of Rous sarcoma virus (RSV) encoding non-myristylated src product with a novel N-terminal domain, results in the immune suppression of challenge tumors induced by RSV. In order to adapt this system to the mouse, we have developed NIH3T3 and Balb/c3T3 (B3T3) cell lines that express 1702src, v-src, c-src, and other src variants, either by transfection or by infection with packaged recombinant Moloney virus (MLV) vectors. The sequence of the 1702 src cDNA, produced by reverse transcription-polymerase chain reaction (RT-PCR), confirmed the previously suggested 1702src N-terminal domain structure, fusing six amino acids from Pr76gag and 39 amino acids of env signal peptide sequence to Ala-76 of src. These cell lines were characterized for src expression and activity, cell compartmentalization of src product, tyrosine phosphorylation substrate specificity and transforming and tumorigenic potential. Based on these parameters, murine cell lines expressing 1702src were characteristically similar to chicken embryo fibroblasts (CEFs) infected with rASV1702. Finally, B3T3/1702src expressors, or membrane fractions of these cells, induced src-specific tumor protection in syngeneic mice against v-src-transformed tumor challenges. Splenic lymphocytes isolated from Balb/c mice inoculated with B3T3/1702src cells showed in vitro cytotoxicity against B3T3/v-src cells but not against untransformed B3T3 cells. Antibodies specific for Lyt2, mouse CD3 and H-2Dd blocked this cytotoxicity, whereas those specific for L3T4 did not, suggesting MHC class I-restricted, CD8-mediated cell killing. These data indicate that Balb/c3T3-expressed 1702src induces a cellular anti-tumor immunity based on src-specific tumor rejection antigens.

3T3 Cells↗