Single-cell transcriptomics reveals multiple chemoresistant properties in leukemic stem and progenitor cells in pediatric AML.

BACKGROUND: Cancer patients can achieve dramatic responses to chemotherapy yet retain resistant tumor cells, which ultimately results in relapse. Although xenograft model studies have identified several cellular and molecular features that are associated with chemoresistance in acute myeloid leukemia (AML), to what extent AML patients exhibit these properties remains largely unknown. RESULTS: We apply single-cell RNA sequencing to paired pre- and post-chemotherapy whole bone marrow samples obtained from 13 pediatric AML patients who had achieved disease remission, and distinguish AML clusters from normal cells based on their unique transcriptomic profiles. Approximately 50% of leukemic stem and progenitor populations actively express leukemia stem cell (LSC) and oxidative phosphorylation (OXPHOS) signatures, respectively. These clusters have a higher chance of tolerating therapy and exhibit an enhanced metabolic program in response to treatment. Interestingly, the transmembrane receptor CD69 is highly expressed in chemoresistant hematopoietic stem cell (HSC)-like populations (named the CD69+ HSC-like subpopulation). Furthermore, overexpression of CD69 results in suppression of the mTOR signaling pathway and promotion of cell quiescence and adhesion in vitro. Finally, the presence of CD69+ HSC-like cells is associated with unfavorable genetic mutations, the persistence of residual tumor cells in chemotherapy, and poor outcomes in independent pediatric and adult public AML cohorts. CONCLUSIONS: Our analysis reveals leukemia stem cell and OXPHOS as two major chemoresistant features in human AML patients. CD69 may serve as a potential biomarker in defining a subpopulation of chemoresistant leukemia stem cells. These findings have important implications for targeting residual chemo-surviving AML cells.

Analyses of Long Noncoding RNA and mRNA Profiles in Subjects with the Phlegm-Dampness Constitution.

BACKGROUND: Constitution in traditional Chinese medicine (TCM) plays a key role in the genesis, development, and prognosis of diseases. Phlegm-dampness constitution (PDC) is one of the nine constitutions in TCM, susceptible to metabolic disorders, which is mainly manifested by profuse phlegm, loose abdomen, and greasy face. Epidemiologic, genomic, and epigenetic studies have been carried out in previous works, confirming that PDC represents a distinctive population with microcosmic changes related to metabolic disorders. However, whether long noncoding RNAs (lncRNAs) play a regulatory role in metabolic disease in subjects with PDC remains largely unknown. We aimed to investigate distinct lncRNA and mRNA expression signatures and lncRNA-mRNA regulatory networks in the phlegm-dampness constitution (PDC). METHODS: The peripheral blood mononuclear cells (PBMCs) were isolated from the subjects with PDC (n = 13) and balanced constitution (BC) (n = 9). The profiles of lncRNAs and mRNAs in PBMCs were analyzed using microarray and further validated with RT-qPCR. Subsequently, pathway analysis was performed to investigate the function of differentially expressed mRNAs by using Ingenuity Pathway Analysis (IPA). RESULTS: Results suggested that some mRNAs, which were regulated by the differentially expressed lncRNAs, were mainly enriched in lipid metabolism and immune inflammation-related pathways. This was consistent with the molecular characteristics of previous studies, indicating that the clinical characteristics of metabolic disorders in PDC might be regulated by lncRNAs. Furthermore, by making coexpression network construction as well as cis-regulated target gene analysis, several lncRNA-mRNA pairs with potential regulatory relationships were identified by bioinformatic analyses, including RP11-317J10.2-CA3, RP11-809C18.3-PIP4K2A, LINC0069-RFTN1, TTTY15-ARHGEF9, and AC135048.13-ORAI3. CONCLUSIONS: This study first revealed that the expression characteristics of lncRNAs/mRNAs may be potential biomarkers, indicating that the distinctive physical and clinical characteristics of PDC might be partially attributed to the specific expression signatures of lncRNAs/mRNAs.

Benzene metabolite hydroquinone promotes DNA homologous recombination repair via the NF-κB pathway.

Benzene, a widespread environmental pollutant, induces DNA double-strand breaks (DSBs) and DNA repair, which may further lead to oncogenic mutations, chromosomal rearrangements and leukemogenesis. However, the molecular mechanisms underlying benzene-induced DNA repair and carcinogenesis remain unclear. The human osteosarcoma cell line (U2OS/DR-GFP), which carries a GFP-based homologous recombination (HR) repair reporter, was treated with hydroquinone, one of the major benzene metabolites, to identify the potential effects of benzene on DSB HR repair. RNA-sequencing was further employed to identify the potential key pathway that contributed to benzene-initiated HR repair. We found that treatment with hydroquinone induced a significant increase in HR. NF-κB pathway, which plays a critical role in carcinogenesis in multiple tumors, was significantly activated in cells recovered from hydroquinone treatment. Furthermore, the upregulation of NF-κB by hydroquinone was also found in human hematopoietic stem and progenitor cells. Notably, the inhibition of NF-κB activity by small molecule inhibitors (QNZ and JSH-23) significantly reduced the frequency of hydroquinone-initiated HR (-1.36- and -1.77-fold, respectively, P < 0.01). Our results demonstrate an important role of NF-κB activity in promoting HR repair induced by hydroquinone. This finding sheds light on the underlying mechanisms involved in benzene-induced genomic instability and leukemogenesis and may contribute to the larger exploration of the influence of other environmental pollutants on carcinogenesis.

Ecological principle meets cancer treatment: treating children with acute myeloid leukemia with low-dose chemotherapy.

Standard chemotherapy regimens for remission induction of pediatric acute myeloid leukemia (AML) are associated with significant morbidity and mortality. We performed a cohort study to determine the impact of reducing the intensity of remission induction chemotherapy on the outcomes of selected children with AML treated with a low-dose induction regimen plus granulocyte colony stimulating factor (G-CSF) (low-dose chemotherapy (LDC)/G-CSF). Complete response (CR) after two induction courses was attained in 87.0% (40/46) of patients receiving LDC/G-CSF. Post-remission therapy was offered to all patients, and included standard consolidation and/or stem cell transplantation. During the study period, an additional 94 consecutive children with AML treated with standard chemotherapy (SDC) for induction (80/94 (85.1%) of the patients attained CR after induction II, P = 0.953) and post-remission. In this non-randomized study, there were no significant differences in 4-year event-free (67.4 vs. 70.7%; P = 0.99) and overall (70.3 vs. 74.6%, P = 0.69) survival in the LDC/G-CSF and SDC cohorts, respectively. After the first course of induction, recovery of white blood cell (WBC) and platelet counts were significantly faster in patients receiving LDC/G-CSF than in those receiving SDC (11.5 vs. 18.5 d for WBCs (P < 0.001); 15.5 vs. 22.0 d for platelets (P < 0.001)). To examine the quality of molecular response, targeted deep sequencing was performed. Of 137 mutations detected at diagnosis in 20 children who attained hematological CR after two courses of LDC/G-CSF (n = 9) or SDC (n = 11), all of the mutations were below the reference value (variant allelic frequency <2.5%) after two courses, irrespective of the treatment group. In conclusion, children with AML receiving LDC/G-CSF appear to have similar outcomes and mutation clearance levels, but significantly lower toxicity than those receiving SDC. Thus, LDC/G-CSF should be further evaluated as an effective alternative to remission induction in pediatric AML.

Chromatin regulator Asxl1 loss and Nf1 haploinsufficiency cooperate to accelerate myeloid malignancy.

ASXL1 is frequently mutated in myeloid malignancies and is known to co-occur with other gene mutations. However, the molecular mechanisms underlying the leukemogenesis associated with ASXL1 and cooperating mutations remain to be elucidated. Here, we report that Asxl1 loss cooperated with haploinsufficiency of Nf1, a negative regulator of the RAS signaling pathway, to accelerate the development of myeloid leukemia in mice. Loss of Asxl1 and Nf1 in hematopoietic stem and progenitor cells resulted in a gain-of-function transcriptional activation of multiple pathways such as MYC, NRAS, and BRD4 that are critical for leukemogenesis. The hyperactive MYC and BRD9 transcription programs were correlated with elevated H3K4 trimethylation at the promoter regions of genes involving these pathways. Furthermore, pharmacological inhibition of both the MAPK pathway and BET bromodomain prevented leukemia initiation and inhibited disease progression in Asxl1Δ/Δ Nf1Δ/Δ mice. Concomitant mutations of ASXL1 and RAS pathway genes were associated with aggressive progression of myeloid malignancies in patients. This study sheds light on the effect of cooperation between epigenetic alterations and signaling pathways on accelerating the progression of myeloid malignancies and provides a rational therapeutic strategy for the treatment of myeloid malignancies with ASXL1 and RAS pathway gene mutations.

PDGFRB mutation and tyrosine kinase inhibitor resistance in Ph-like acute lymphoblastic leukemia.

Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) comprises ∼10% to 15% of childhood ALL cases, many of which respond exquisitely to tyrosine kinase inhibitors (TKIs), for example, imatinib in PDGFRB-rearranged ALL. However, some cases developed drug resistance to TKIs and the mechanisms are poorly understood. In this study, we identified a novel PDGFRB fusion gene, namely AGGF1-PDGFRB, and functionally characterized its oncogenic potential in vitro. Further genomic profiling of longitudinally collected samples during treatment revealed the emergence of a mutation, PDGFRBC843G , which directly conferred resistance to all generations of ABL TKIs, including imatinib, dasatinib, nilotinib, and ponatinib. PDGFRB-mutant leukemia cells are highly sensitive to multitarget kinase inhibitor CHZ868, suggesting potential therapeutic options for some patients resistant to ABL TKIs. In summary, we describe a complex clonal evolution pattern in Ph-like ALL and identified a novel PDGFRB point mutation that drives leukemia relapse after ABL TKI treatment.

Loss of Asxl2 leads to myeloid malignancies in mice.

ASXL2 is frequently mutated in acute myeloid leukaemia patients with t(8;21). However, the roles of ASXL2 in normal haematopoiesis and the pathogenesis of myeloid malignancies remain unknown. Here we show that deletion of Asxl2 in mice leads to the development of myelodysplastic syndrome (MDS)-like disease. Asxl2-/- mice have an increased bone marrow (BM) long-term haematopoietic stem cells (HSCs) and granulocyte-macrophage progenitors compared with wild-type controls. Recipients transplanted with Asxl2-/- and Asxl2+/- BM cells have shortened lifespan due to the development of MDS-like disease or myeloid leukaemia. Paired daughter cell assays demonstrate that Asxl2 loss enhances the self-renewal of HSCs. Deletion of Asxl2 alters the expression of genes critical for HSC self-renewal, differentiation and apoptosis in Lin-cKit+ cells. The altered gene expression is associated with dysregulated H3K27ac and H3K4me1/2. Our study demonstrates that ASXL2 functions as a tumour suppressor to maintain normal HSC function.

Study of the water transportation characteristics of marsh saline soil in the Yellow River Delta.

One-dimensional soil column water infiltration and capillary adsorption water tests were conducted in the laboratory to study the water transportation characteristics of marsh saline soil in the Yellow River Delta, providing a theoretical basis for the improvement, utilization and conservation of marsh saline soil. The results indicated the following: (1) For soils with different vegetation covers, the cumulative infiltration capacity increased with the depth of the soil layers. The initial infiltration rate of soils covered by Suaeda and Tamarix chinensis increased with depth of the soil layers, but that of bare soil decreased with soil depth. (2) The initial rate of capillary rise of soils with different vegetation covers showed an increasing trend from the surface toward the deeper layers, but this pattern with respect to soil depth was relatively weak. (3) The initial rates of capillary rise were lower than the initial infiltration rates, but infiltration rate decreased more rapidly than capillary water adsorption rate. (4) The two-parameter Kostiakov model can very well-simulate the changes in the infiltration and capillary rise rates of wetland saline soil. The model simulated the capillary rise rate better than it simulated the infiltration rate. (5) There were strong linear relationships between accumulative infiltration capacity, wetting front, accumulative capillary adsorbed water volume and capillary height.

Temperature and moisture responses to carbon mineralization in the biochar-amended saline soil.

This study assessed the effects of temperature and moisture on carbon mineralization (Cmin) in a saline soil system with biochar amendment. The dynamics of Cmin were monitored in a biochar-amended saline soil for 220days by incubation experiments under different conditions of temperature (15°C, 25°C and 35°C) and moisture (30%, 70% and 105% of the water-holding capacity). Results showed that as the incubation temperature rose, cumulative Cmin consistently increased in soil added with 0-4% biochar. The two-compartment model could well describe the dynamics of Cmin. The temperature rise increased the concentration of labile C in soil, but reduced the turnover time of labile and recalcitrant C pools and the value of temperature coefficient Q10. The response of Cmin to moisture was varying in soil amended with different levels of biochar. In the control treatment (soil alone), cumulative Cmin increased only when soil moisture was >105%. In the biochar treatments, however, 70% of water-holding capacity was optimal for Cmin, except for 2%-biochar treatment at 35°C. The findings highlight the necessity to consider the combined effects of soil moisture, temperature and the amount of biochar added for assessing Cmin in biochar-amended saline soils.

Loss of Asxl1 Alters Self-Renewal and Cell Fate of Bone Marrow Stromal Cell, Leading to Bohring-Opitz-like Syndrome in Mice.

De novo ASXL1 mutations are found in patients with Bohring-Opitz syndrome, a disease with severe developmental defects and early childhood mortality. The underlying pathologic mechanisms remain largely unknown. Using Asxl1-targeted murine models, we found that Asxl1 global loss as well as conditional deletion in osteoblasts and their progenitors led to significant bone loss and a markedly decreased number of bone marrow stromal cells (BMSCs) compared with wild-type littermates. Asxl1(-/-) BMSCs displayed impaired self-renewal and skewed differentiation, away from osteoblasts and favoring adipocytes. RNA-sequencing analysis revealed altered expression of genes involved in cell proliferation, skeletal development, and morphogenesis. Furthermore, gene set enrichment analysis showed decreased expression of stem cell self-renewal gene signature, suggesting a role of Asxl1 in regulating the stemness of BMSCs. Importantly, re-introduction of Asxl1 normalized NANOG and OCT4 expression and restored the self-renewal capacity of Asxl1(-/-) BMSCs. Our study unveils a pivotal role of ASXL1 in the maintenance of BMSC functions and skeletal development.

An integrated use of topography with RSI in gully mapping, Shandong Peninsula, China.

Taking the Quickbird optical satellite imagery of the small watershed of Beiyanzigou valley of Qixia city, Shandong province, as the study data, we proposed a new method by using a fused image of topography with remote sensing imagery (RSI) to achieve a high precision interpretation of gully edge lines. The technique first transformed remote sensing imagery into HSV color space from RGB color space. Then the slope threshold values of gully edge line and gully thalweg were gained through field survey and the slope data were segmented using thresholding, respectively. Based on the fused image in combination with gully thalweg thresholding vectors, the gully thalweg thresholding vectors were amended. Lastly, the gully edge line might be interpreted based on the amended gully thalweg vectors, fused image, gully edge line thresholding vectors, and slope data. A testing region was selected in the study area to assess the accuracy. Then accuracy assessment of the gully information interpreted by both interpreting remote sensing imagery only and the fused image was performed using the deviation, kappa coefficient, and overall accuracy of error matrix. Compared with interpreting remote sensing imagery only, the overall accuracy and kappa coefficient are increased by 24.080% and 264.364%, respectively. The average deviations of gully head and gully edge line are reduced by 60.448% and 67.406%, respectively. The test results show the thematic and the positional accuracy of gully interpreted by new method are significantly higher. Finally, the error sources for interpretation accuracy by the two methods were analyzed.

Rictor/mammalian target of rapamycin 2 regulates the development of Notch1 induced murine T-cell acute lymphoblastic leukemia via forkhead box O3.

Mammalian target of rapamycin (mTOR) is composed of two distinct biochemical complexes, mTORC1 and mTORC2. In response to nutrients and growth factors, mTORC1 is known to control cellular growth by regulating the translational regulators S6 kinase 1 and 4E binding protein 1, whereas mTORC2 mediates cell proliferation and survival by activating Akt through phosphorylation at Ser473. Studies have shown that the deregulation of mTORC2 leads to the development of myeloproliferative disorder and leukemia in the phosphatase and tensin homolog deleted on chromosome ten (PTEN)-deleted mouse model. However, the mechanism by which mTORC2 specifically affects leukemogenesis is still not fully understood. Here, we investigated the role of mTORC2 in NOTCH1-driven T-cell acute lymphoblastic leukemia (T-ALL) in a Rictor-deficient mouse model. We found that, by deleting Rictor, an essential component of mTORC2, leukemia progression was significantly suppressed by arresting a greater proportion of Rictor(△/△) leukemic cells at the G0 phase of the cell cycle. Furthermore, the absence of Rictor led to the overexpression of chemotaxis-related genes, such as CCR2, CCR4 and CXCR4, which contributed to the homing and migration of Rictor-deficient T-ALL cells to the spleen but not the bone marrow. In addition, we demonstrated that inactivation of mTORC2 caused the overexpression of forkhead box O3 and its downstream effectors and eased the progression of leukemia in T-ALL mice. Our study thus indicates that forkhead box O3 could be a potential drug target for the treatment of T-ALL leukemia.

A quantitative study of gully erosion based on object-oriented analysis techniques: a case study in Beiyanzikou catchment of Qixia, Shandong, China.

This paper took a subregion in a small watershed gully system at Beiyanzikou catchment of Qixia, China, as a study and, using object-orientated image analysis (OBIA), extracted shoulder line of gullies from high spatial resolution digital orthophoto map (DOM) aerial photographs. Next, it proposed an accuracy assessment method based on the adjacent distance between the boundary classified by remote sensing and points measured by RTK-GPS along the shoulder line of gullies. Finally, the original surface was fitted using linear regression in accordance with the elevation of two extracted edges of experimental gullies, named Gully 1 and Gully 2, and the erosion volume was calculated. The results indicate that OBIA can effectively extract information of gullies; average range difference between points field measured along the edge of gullies and classified boundary is 0.3166 m, with variance of 0.2116 m. The erosion area and volume of two gullies are 2141.6250 m(2), 5074.1790 m(3) and 1316.1250 m(2), 1591.5784 m(3), respectively. The results of the study provide a new method for the quantitative study of small gully erosion.

Identification of functional cooperative mutations of SETD2 in human acute leukemia.

Acute leukemia characterized by chromosomal rearrangements requires additional molecular disruptions to develop into full-blown malignancy, yet the cooperative mechanisms remain elusive. Using whole-genome sequencing of a pair of monozygotic twins discordant for MLL (also called KMT2A) gene-rearranged leukemia, we identified a transforming MLL-NRIP3 fusion gene and biallelic mutations in SETD2 (encoding a histone H3K36 methyltransferase). Moreover, loss-of-function point mutations in SETD2 were recurrent (6.2%) in 241 patients with acute leukemia and were associated with multiple major chromosomal aberrations. We observed a global loss of H3K36 trimethylation (H3K36me3) in leukemic blasts with mutations in SETD2. In the presence of a genetic lesion, downregulation of SETD2 contributed to both initiation and progression during leukemia development by promoting the self-renewal potential of leukemia stem cells. Therefore, our study provides compelling evidence for SETD2 as a new tumor suppressor. Disruption of the SETD2-H3K36me3 pathway is a distinct epigenetic mechanism for leukemia development.

Downregulation of RUNX1/CBFß by MLL fusion proteins enhances hematopoietic stem cell self-renewal.

RUNX1/CBFß (core binding factor [CBF]) is a heterodimeric transcription factor complex that is frequently involved in chromosomal translocations, point mutations, or deletions in acute leukemia. The mixed lineage leukemia (MLL) gene is also frequently involved in chromosomal translocations or partial tandem duplication in acute leukemia. The MLL protein interacts with RUNX1 and prevents RUNX1 from ubiquitin-mediated degradation. RUNX1/CBFß recruits MLL to regulate downstream target genes. However, the functional consequence of MLL fusions on RUNX1/CBFß activity has not been fully understood. In this report, we show that MLL fusion proteins and the N-terminal MLL portion of MLL fusions downregulate RUNX1 and CBFß protein expression via the MLL CXXC domain and flanking regions. We confirmed this finding in Mll-Af9 knock-in mice and human M4/M5 acute myeloid leukemia (AML) cell lines, with or without MLL translocations, showing that MLL translocations cause a hypomorph phenotype of RUNX1/CBFß. Overexpression of RUNX1 inhibits the development of AML in Mll-Af9 knock-in mice; conversely, further reducing Runx1/Cbfß levels accelerates MLL-AF9-mediated AML in bone marrow transplantation assays. These data reveal a newly defined negative regulation of RUNX1/CBFß by MLL fusion proteins and suggest that targeting RUNX1/CBFß levels may be a potential therapy for MLLs.

Gene and genome parameters of mammalian liver circadian genes (LCGs).

The mammalian circadian system controls various physiology processes and behavior responses by regulating thousands of circadian genes with rhythmic expressions. In this study, we redefined circadian-regulated genes based on published results in the mouse liver and compared them with other gene groups defined relative to circadian regulations, especially the non-circadian-regulated genes expressed in liver at multiple molecular levels from gene position to protein expression based on integrative analyses of different datasets from the literature. Based on the intra-tissue analysis, the liver circadian genes or LCGs show unique features when compared to other gene groups. First, LCGs in general have less neighboring genes and larger in both genomic and 3'-UTR lengths but shorter in CDS (coding sequence) lengths. Second, LCGs have higher mRNA and protein abundance, higher temporal expression variations, and shorter mRNA half-life. Third, more than 60% of LCGs form major co-expression clusters centered in four temporal windows: dawn, day, dusk, and night. In addition, larger and smaller LCGs are found mainly expressed in the day and night temporal windows, respectively, and we believe that LCGs are well-partitioned into the gene expression regulatory network that takes advantage of gene size, expression constraint, and chromosomal architecture. Based on inter-tissue analysis, more than half of LCGs are ubiquitously expressed in multiple tissues but only show rhythmical expression in one or limited number of tissues. LCGs show at least three-fold lower expression variations across the temporal windows than those among different tissues, and this observation suggests that temporal expression variations regulated by the circadian system is relatively subtle as compared with the tissue expression variations formed during development. Taken together, we suggest that the circadian system selects gene parameters in a cost effective way to improve tissue-specific functions by adapting temporal variations from the environment over evolutionary time scales.

MLL fusion proteins preferentially regulate a subset of wild-type MLL target genes in the leukemic genome.

MLL encodes a histone methyltransferase that is critical in maintaining gene expression during embryonic development and hematopoiesis. 11q23 translocations result in the formation of chimeric MLL fusion proteins that act as potent drivers of acute leukemia. However, it remains unclear what portion of the leukemic genome is under the direct control of MLL fusions. By comparing patient-derived leukemic cell lines, we find that MLL fusion-bound genes are a small subset of that recognized by wild-type MLL. In an inducible MLL-ENL model, MLL fusion protein binding and changes in H3K79 methylation are limited to a specific portion of the genome, whereas wild-type MLL distributes to a much larger set of gene loci. Surprisingly, among 223 MLL-ENL-bound genes, only 12 demonstrate a significant increase in mRNA expression on induction of the fusion protein. In addition to Hoxa9 and Meis1, this includes Eya1 and Six1, which comprise a heterodimeric transcription factor important in several developmental pathways. We show that Eya1 has the capacity to immortalize hematopoietic progenitor cells in vitro and collaborates with Six1 in hematopoietic transformation assays. Altogether, our data suggest that MLL fusions contribute to the development of acute leukemia through direct activation of a small set of target genes.

A novel role for minimal introns: routing mRNAs to the cytosol.

BACKGROUND: Introns and their splicing are tightly coupled with the subsequent mRNA maturation steps, especially nucleocytoplasmic export. A remarkable fraction of vertebrate introns have a minimal size of about 100 bp, while majority of introns expand to several kilobases even megabases in length. PRINCIPAL FINDINGS: We carried out analyses on the evolution and function of minimal introns (50-150 bp) in human and mouse genomes. We found that minimal introns are conserved in terms of both length and sequence. They are preferentially located toward 3' end of mRNA and non-randomly distributed among chromosomes. Both the evolutionary conservation and non-random distribution are indicative of biological relevance. We showed that genes with minimal introns have higher abundance, larger size, and tend to be universally expressed as compared to genes with only large introns and intron-less genes. Genes with minimal introns replicate earlier and preferentially reside in the vicinities of open chromatin, suggesting their unique nuclear position and potential relevance to the regulation of gene expression and transcript export. CONCLUSIONS: Based on these observations, we proposed a nuclear-export routing model, where minimal introns play a regulatory role in selectively exporting the highly abundant and large housekeeping genes that reside at the surface of chromatin territories, and thus preventing entanglement with other genes located at the interior locations.

How do variable substitution rates influence Ka and Ks calculations?

The ratio of nonsynonymous substitution rate (Ka) to synonymous substitution rate (Ks) is widely used as an indicator of selective pressure at sequence level among different species, and diverse mutation models have been incorporated into several computing methods. We have previously developed a new gamma-MYN method by capturing a key dynamic evolution trait of DNA nucleotide sequences, in consideration of varying mutation rates across sites. We now report a further improvement of NG, LWL, MLWL, LPB, MLPB, and YN methods based on an introduction of gamma distribution to illustrate the variation of raw mutation rate over sites. The novelty comes in two ways: (1) we incorporate an optimal gamma distribution shape parameter a into gamma-NG, gamma-LWL, gamma-MLWL, gamma-LPB, gamma-MLPB, and gamma-YN methods; (2) we investigate how variable substitution rates affect the methods that adopt different models as well as the interplay among four evolutional features with respect to Ka/Ks computations. Our results suggest that variable substitution rates over sites under negative selection exhibit an opposite effect on omega estimates compared with those under positive selection. We believe that the sensitivity of our new methods has been improved than that of their original methods under diverse conditions and it is advantageous to introduce novel parameters for Ka/Ks computation.

On the nature of human housekeeping genes.

Using a collection of expressed sequence tag (EST) data, we re-evaluated the correlation of tissue specificity with genomic structure, phyletic age, evolutionary rate and promoter architecture of human genes. We found that housekeeping genes are less compact and older than tissue-specific genes, and they evolve more slowly in terms of both coding and core promoter sequences. Housekeeping genes primarily use CpG-dependent core promoters, whereas the majority of tissue-specific genes possess neither CpG-islands nor TATA-boxes in their core promoters.