Biomedical subjects

Guojun Yang

Publications and source records attributed to Guojun Yang.

12 recordsLinked to original sources

PubMed2006

Dramatic amplification of a rice transposable element during recent domestication.

Despite the prevalence of transposable elements in the genomes of higher eukaryotes, what is virtually unknown is how they amplify to very high copy numbers without killing their host. Here, we report the discovery of rice strains where a miniature inverted-repeat transposable element (mPing) has amplified from approximately 50 to approximately 1,000 copies in four rice strains. We characterized 280 of the insertions and found that 70% were within 5 kb of coding regions but that insertions into exons and introns were significantly underrepresented. Further analyses of gene expression and transposable-element activity demonstrate that the ability of mPing to attain high copy numbers is because of three factors: (i) the rapid selection against detrimental insertions, (ii) the neutral or minimal effect of the remaining insertions on gene transcription, and (iii) the continued mobility of mPingelements in strains that already have > 1,000 copies. The rapid increase in mPing copy number documented in this study represents a potentially valuable source of population diversity in self-fertilizing plants like rice.

Base Sequence↗

PubMed2006

A rice Tc1/mariner-like element transposes in yeast.

The Tc1/mariner transposable element superfamily is widely distributed in animal and plant genomes. However, no active plant element has been previously identified. Nearly identical copies of a rice (Oryza sativa) Tc1/mariner element called Osmar5 in the genome suggested potential activity. Previous studies revealed that Osmar5 encoded a protein that bound specifically to its own ends. In this report, we show that Osmar5 is an active transposable element by demonstrating that expression of its coding sequence in yeast promotes the excision of a nonautonomous Osmar5 element located in a reporter construct. Element excision produces transposon footprints, whereas element reinsertion occurs at TA dinucleotides that were either tightly linked or unlinked to the excision site. Several site-directed mutations in the transposase abolished activity, whereas mutations in the transposase binding site prevented transposition of the nonautonomous element from the reporter construct. This report of an active plant Tc1/mariner in yeast will provide a foundation for future comparative analyses of animal and plant elements in addition to making a new wide host range transposable element available for plant gene tagging.

Base Sequence↗

PubMed2006

Combined IL-21 and low-dose IL-2 therapy induces anti-tumor immunity and long-term curative effects in a murine melanoma tumor model.

BACKGROUND: In vivo studies have recently demonstrated that interleukin 21 (IL-21) enhances the anti-tumor function of T-cells and NK cells in murine tumor models, and the combined use of IL-21 and IL-15 has resulted in prolonged tumor regression and survival in mice with previously established tumors. However, the combined anti-tumor effects of IL-21 and low dose IL-2 have not been studied even though IL-2 has been approved for human use, and, at low dose administration, stimulates the proliferation of memory T cells, and does not significantly increase antigen-induced apoptosis or regulatory T cell (Treg) expansion. This study examined whether recombinant IL-21 alone or in combination with low-dose IL-2 could improve the in vivo anti-tumor function of naïve, tumor-antigen specific CD8+ T cells in a gp100(25-33) T cell receptor transgenic pmel murine melanoma model. METHODS: Congenic C57BL/6 (Ly5.2) mice bearing subcutaneous B16F10 melanoma tumors were sublethally irradiated to induce lymphopenia. After irradiation naive pmel splenocytes were adoptively transferred, and mice were immunized with bone marrow-derived dendritic cells pulsed with human gp100(25-33) (hgp100(25-33)). Seven days after vaccination groups of mice received 5 consecutive days of intraperitoneal administration of IL-2 alone (20 x 10(3) IU), IL-21 alone (20 microg) or IL-21 and IL-2. Control animals received no cytokine therapy. RESULTS: IL-21 alone and IL-2 alone both delayed tumor progression, but only IL-21 significantly augmented long-term survival (20%) compared to the control group. However, combination therapy with IL-21 and IL-2 resulted in the highest long-term (>150 days) tumor-free survival frequency of 46%. Animals that were tumor-free for > 150 days demonstrated tumor-specific protection after rechallenge with B16F10 melanoma cells. At peak expansion (21 days post vaccination), the combination of IL-21 plus IL-2 resulted in a 2- to 3-fold higher absolute number of circulating tumor antigen-specific pmel CD8+ T cells than was stimulated by IL-2 or IL-21 alone. Pmel CD8+ T cells were predominantly partitioned into central memory (CD62L+/CD127+) or effector-memory (CD62L-/CD127+) phenotypes by day 28-post vaccination in IL-21 + IL-2 treated mice. CONCLUSION: These observations support the potential use of IL-21 and low-dose IL-2 therapy in combination with a tumor-antigen vaccine and lymphopenic conditioning in future cancer clinical trials to maintain high numbers of anti-tumor memory CD8+ T cells with the potential to sustain long term tumor regression and survival.

Journal Article↗

PubMed2005

HideNseek, a post-genome approach to locate transgenes exemplified in Arabidopsis thaliana.

SUMMARY: Determination of transgene location is essential for investigating the effects of position on transgene expression levels and facilitates cloning of the resident gene affected by insertion. Currently used PCR-based approaches for determination of transgene location are relatively complicated and often fail when the transgene is duplicated, rearranged or fragmented. HideNseek is a new bioinformatics tool that allows computation of transgene locations, provided that a suitable genomic restriction enzyme digestion profile is available. Since the new approach is not based on the terminal sequences of the transgene insert, it is less sensitive to transgene duplication, rearrangement or fragmentation. HideNseek has been tested experimentally and by in silico simulation. The experimental example provided here shows that this simple approach is feasible, permitting rapid location of transgenes with little bench work. AVAILABILITY: available on request from the authors. SUPPLEMENTARY DATA: HideNseek input and output examples, experimental procedures and figures showing experimental results are provided as supplementary files: Supplementary material 1, 2, 3 and Supplementary figures (Figs 1 and 2), respectively. Supplementary data is available at Bioinformatics online.

Algorithms↗

PubMed2005

Interleukin-7-dependent expansion and persistence of melanoma-specific T cells in lymphodepleted mice lead to tumor regression and editing.

Active-specific immunotherapy with dendritic cells loaded with peptide derived from the melanoma antigen, gp100, failed to mediate regression of established B16F10 melanoma in normal mice. Dendritic cell vaccination induced activation and subsequent deletion of adoptively transferred naive CD8+ T-cell receptor transgenic (pmel-1) T cells specific for gp100 in normal mice. In lymphodepleted mice, dendritic cell vaccination produced greater T-cell expansion, long-term persistence of memory T cells, and tumor regression. Most tumors that persisted in the presence of functional memory T cells had either lost or exhibited reduced expression of MHC class I or gp100 proteins. In contrast to other naive T cells, pmel-1 T cells adoptively transferred to lymphodepleted mice exhibited faster proliferation and a more differentiated phenotype after exposure to peptide-pulsed dendritic cells. Proliferation and persistence of pmel-1 T cells was highly dependent on interleukin-7 (IL-7) in irradiated mice, and IL-15 when IL-7 was neutralized, two critical homeostatic cytokines produced in response to the irradiation-induced lymphodepletion.

Animals↗

PubMed2005

A two-edged role for the transposable element Kiddo in the rice ubiquitin2 promoter.

Miniature inverted repeat transposable elements (MITEs) are thought to be a driving force for genome evolution. Although numerous MITEs are found associated with genes, little is known about their function in gene regulation. Whereas the rice ubiquitin2 (rubq2) promoter in rice (Oryza sativa) line IR24 contains two nested MITEs (Kiddo and MDM1), that in line T309 has lost Kiddo, providing an opportunity to understand the role of MITEs in promoter function. No difference in endogenous rubq2 transcript levels between T309 and IR24 was evident using RT-PCR. However, promoter analysis using both transient and stably transformed calli revealed that Kiddo contributed some 20% of the total expression. Bisulfite genomic sequencing of the rubq2 promoters revealed specific DNA methylation at both symmetric and asymmetric cytosine residues on the MITE sequences, possibly induced by low levels of homologous transcripts. When methylation of the MITEs was blocked by 5-azacytidine treatment, a threefold increase in the endogenous rubq2 transcript level was detected in IR24 compared with that in T309. Together with the observed MITE methylation pattern, the detection of low levels of transcripts, but not small RNAs, corresponding to Kiddo and MDM1 suggested that RNA-dependent DNA methylation is induced by MITE transcripts. We conclude that, although Kiddo enhances transcription from the rubq2 promoter, this effect is mitigated by sequence-specific epigenetic modification.

Base Sequence↗

PubMed2005

Organization, not duplication, triggers silencing in a complex transgene locus in rice.

Despite the presence in nature of many functional gene families that contain several to many highly similar sequences, the presence of identical DNA sequence repeats is widely thought to predispose transgene inserts to homology dependent gene silencing (HDGS). The induction of transcriptional gene silencing (TGS) by RNAs homologous to promoter sequences has been reported recently in Arabidopsis and humans. However, mechanisms for TGS have not been studied in detail for rice, the most widely cultivated crop plant. Taking advantage of a well-characterized homozygous silenced transgenic rice line (siJKA), supertransformation was performed with a binary vector bearing mUbi1 and 35S promoter sequences identical to those in the resident transgenes. Analysis of the incoming and resident transgenes in the supertransformants revealed that the incoming mUbi1 transgene promoter was not silenced whereas the incoming 35S transgene promoter was silenced. That the resident silenced mUbi1-bar was not reactivated in these experiments as a result of passage through tissue culture and regeneration was established by the finding that regenerants from siJKA immature embryos were all silenced for mUbi1-bar. In a parallel experiment, when wild type rice calli were transformed with the same binary vector, neither of the incoming transgene promoters was silenced. Following 5-azacytidine (5-azaC) treatment of siJKA, aberrant RNA species corresponding to the 35S promoter, but not to the mUbi1 promoter, were detected. Nevertheless, no 21-25 nt RNAs corresponding to the 35S promoter sequence were detected. These results, together with detailed analyses of the progenies from the primary transformants and supertransformants, revealed that HDGS of the resident silenced locus was caused not by simple transgene duplication, but by aberrant transcripts derived from rearranged promoters present in siJKA. Practical consequences of this study include a justification for the use of multiple copies of a given promoter for transformation without inducing silencing, provided that their genomic integration does not result in aberrant transcription of the promoters.

Azacitidine↗

PubMed2004

Functional impairment of CD8(+) T cells by regulatory T cells during persistent retroviral infection.

The establishment of viral persistence generally requires evasion of the host CD8(+) T cell response. Here we describe a form of evasion wherein the CD8(+) T cells are fully capable of recognizing their cognate antigen but their effector functions are suppressed by regulatory T cells. Virus-specific CD8(+) T cells adoptively transferred into mice persistently infected with Friend virus proliferated and appeared activated, but failed to produce IFNgamma or reduce virus loads. Cotransfer experiments revealed that a subpopulation of CD4(+) T cells from persistently infected mice suppressed IFNgamma production by the CD8(+) T cells. Treatment of persistently infected mice with anti-GITR antibody to ameliorate suppression by regulatory T cells significantly improved IFNgamma production by transferred CD8(+) T cells and allowed a significant reduction in viral loads. The results indicate that CD4(+) regulatory T cells contribute to viral persistence and demonstrate an immunotherapy for treating chronic retroviral infections.

Acute Disease↗

PubMed2004

Reduction of retrovirus-induced immunosuppression by in vivo modulation of T cells during acute infection.

Chronic infection with Friend retrovirus is associated with suppressed antitumor immune responses. In the present study we investigated whether modulation of T-cell responses during acute infection would restore antitumor immunity in persistently infected mice. T-cell modulation was done by treatments with DTA-1 anti- glucocorticoid-induced tumor necrosis factor receptor monoclonal antibodies. The DTA-1 monoclonal antibody is nondepleting and delivers costimulatory signals that both enhance the activation of effector T cells and inhibit suppression by regulatory T cells. DTA-1 therapy produced faster Th1 immune responses, significant reductions in both acute virus loads and pathology and, most importantly, long-term improvement of CD8(+) T-cell-mediated antitumor responses.

Acute Disease↗

PubMed2003

MAK, a computational tool kit for automated MITE analysis.

Miniature inverted repeat transposable elements (MITEs) are ubiquitous and numerous in higher eukaryotic genomes. Analysis of MITE families is laborious and time consuming, especially when multiple MITE families are involved in the study. Based on the structural characteristics of MITEs and genetic principles for transposable elements (TEs), we have developed a computational tool kit named MITE analysis kit (MAK) to automate the processes (http://perl.idmb.tamu.edu/mak.htm). In addition to its ability to routinely retrieve family member sequences and to report the positions of these elements relative to the closest neighboring genes, MAK is a powerful tool for revealing anchor elements that link MITE families to known transposable element families. Implementation of the MAK is described, as are genetic principles and algorithms used in its derivation. Test runs of the programs for several MITE families yielded anchor sequences that retain TIRs and coding regions reminiscent of transposases. These anchor sequences are consistent with previously reported putative autonomous elements for these MITE families. Furthermore, analysis of two MITE families with no known links to any transposon family revealed two novel transposon families, namely Math and Kid, belonging to the IS5/Harbinger/PIF superfamily.

Animals↗

PubMed2003

MDM-1 and MDM-2: two mutator-derived MITE families in rice.

Numerous miniature inverted repeat transposable elements (MITEs) are present in the rice genome but their transposition mechanisms are unknown. In this report, we present evidence that two novel MITE families may have arisen from Mutator-related transposable elements and thus may use a transposition mechanism similar to that of Mutator elements. Two families of novel MITEs, namely, MDM-1 and MDM-2, were identified by searching for MITEs nested with Kiddo, a previously identified MITE family. MDM-1 and MDM-2 bear hallmarks of Mutator elements, such as long terminal inverted repeats (LTIRs), 9-bp target-site duplications (TSDs), and putative transposase binding sites. Strikingly, the MDM-1 family has a 9-bp terminus identical to that of a rice Mutator-like element ( MULE-9) and the MDM-2 family has an 8-bp terminus identical to that of the maize autonomous Mutator element MuDR. A putative transposase homologous to MURA protein is identified for the MDM-2 family. Thus, these two novel MITE families, with a total copy number of several hundred in rice, are designated Mutator-derived MITEs ( MDMs). Interestingly, sequence decay analysis of MDM families revealed a number of insertion site duplications (ISDs) in the alignment gaps, and widespread historical nesting events are proposed to account for the existence of these ISDs. In addition to its value for discovering new MITEs, the nesting analysis approach used in this study simultaneously identifies MITE insertion polymorphisms.

Base Sequence↗

PubMedDate not supplied

Efficacy of GM-CSF-producing tumor vaccine after docetaxel chemotherapy in mice bearing established Lewis lung carcinoma.

In this report, we evaluated the efficacy of a GM-CSF-producing tumor vaccine given before and after docetaxel in mice bearing established lung tumors. Mice bearing established 3LL tumors were treated with docetaxel and tumor vaccines transduced with either control or GM-CSF adenoviral vectors. Docetaxel (5-20 mg/kg) treatment alone had only a minimal effect on growth of established 3LL tumors in vivo, although docetaxel was cytotoxic to 3LL cells in vitro. When mice bearing established 3LL tumors were pretreated with docetaxel followed by vaccination with irradiated GM-CSF- transduced 3LL tumor cells, significant tumor regression and prolonged survival were observed compared with chemotherapy alone. Delaying docetaxel treatment until after tumor vaccination abrogated the vaccine's anti-tumor effects. Mice that survived treatment were able to resist a lethal rechallenge of 3LL tumor cells. Memory CTL specific for an epitope (MUT-1) derived from 3LL were detected in surviving mice. Docetaxel induced a mild lymphodepletion in mice, both CD4 and CD8 subsets were reduced in LN and spleens. Interestingly, docetaxel also diminished the number of memory CD8+ T cells (CD122+) and possible CD4+ CD25+ Foxp3+ natural Treg cells. Docetaxel treatment did not affect antigen-driven proliferation of naive T cells but significantly promoted survival of activated T cells. Thus, augmentation of vaccine induced antitumor immunity in docetaxel-treated mice primarily due to the enhanced survival of antigen-experienced T cells.

Adenoviridae↗