Biology subjects

Akgul, B.

Publications and source records attributed to Akgul, B..

7 recordsLinked to original sources

bioRxiv2026

METTL3 modulates cell viability and motility in HCC1143 and MDA-MB-231 triple-negative breast cancer cells

The m6A methyltransferase METTL3 functions as a critical oncogenic driver in triple-negative breast cancer (TNBC). However, its specific downstream targets and mechanistic functions in less metastatic TNBC subtypes remain poorly characterized. To address this, we evaluated METTL3 expression and the phenotypic effects of its siRNA-mediated knockdown in normal mammary epithelial (MCF10A), low-metastatic TNBC (HCC1143), and high-metastatic TNBC (MDA-MB-231) cell lines. We assessed global m6A levels, cell viability, cell cycle progression, and migration. To uncover specific downstream pathways, transcriptomic profiling was performed on HCC1143 cells, followed by RT-qPCR validation and m6A site prediction. METTL3 depletion reduced global m6A levels and cell viability across all cell lines. Notably, in low-metastatic HCC1143 cells, METTL3 knockdown induced a pronounced G2/M cell cycle arrest and dramatically impaired migratory capacity. Transcriptomic analysis of HCC1143 revealed altered expression of genes associated with the observed phenotypic changes. Specifically, critical transcripts harboring predicted m6A motifs, including LIMK1, CCNB2, and CDH1, were significantly dysregulated, pointing to potential alterations in pathways governing cytoskeletal remodeling, actin organization, and cell-cell adhesion. Taken together, we propose that METTL3 promotes cell viability and motility in low-metastatic TNBC by regulating key transcripts involved in cell cycle progression and actin dynamics. Significance StatementEpitranscriptomic studies on TNBC predominantly focus on highly metastatic models, leaving less aggressive subtypes poorly understood. This study uniquely addresses this gap by investigating the function of METTL3 in HCC1143, a low-metastatic TNBC cell line, alongside aggressive TNBC cell lines. We discovered that METTL3 depletion uniquely triggers a severe halt in cell division (G2/M arrest) in HCC1143 cells, while universally disrupting actin-associated cell motility across different backgrounds. These findings demonstrate that METTL3 acts as a context-dependent modulator of cell fate rather than a monolithic driver. Ultimately, highlighting these distinct cellular responses underscores the need to consider specific molecular backgrounds when evaluating epitranscriptomic targets in heterogeneous cancers, such as TNBC.

cancer biology↗

bioRxiv2023

Expression patterns of m6A RNA methylation regulators under apoptotic conditions in various human cancer cell lines

The emerging evidence suggests that epitranscriptomics changes play a crucial role in the pathogenesis of cancer. However, expression patterns of m6A RNA modifiers under apoptotic conditions are unknown. We measured the transcript abundance of m6A RNA modifiers under cisplatin- and tumor necrosis factor alpha (TNF-)-induced apoptotic conditions. In general, the abundance of m6A modifiers is increased upon cisplatin treatment whereas TNF- treatment has led to a reduction in their expression. Specifically, cisplatin-induced apoptosis, but not TNF- mediated apoptosis, lowered the abundance of METTL14 and FTO transcripts. Additionally, cisplatin treatment plummeted the abundance of IGF2BP2 and IGF2BP3 readers following cisplatin treatment. These results suggest that differential response of cancer cells to apoptotic inducers could be partially due to m6A RNA modifiers.

cancer biology↗

bioRxiv2023

Epitranscriptomics m6A analyses reveal distinct m6A marks under tumor necrosis α (TNF-α)-induced apoptotic conditions in HeLa cells

TNF- is a ligand that induces both intrinsic and extrinsic apoptotic pathways in HeLa cells by modulating complex gene regulatory mechanisms. However, the full spectrum of TNF--modulated epitranscriptomic m6A marks is unknown. We employed a genomewide approach to examine the extent of m6A RNA modifications under TNF--modulated apoptotic conditions in HeLa cells. miCLIP-seq analyses revealed a plethora of m6A marks on 632 target mRNAs with an enrichment on 99 mRNAs associated with apoptosis. Interestingly, the m6A RNA modification patterns were quite different under cisplatin- and TNF--mediated apoptotic conditions. We then examined the abundance and translational efficiencies of several mRNAs under METTL3 knockdown and/or TNF- treatment conditions. Our analyses showed changes in the translational efficiency of TP53INP1 mRNA based on the polysome profile analyses. Additionally, TP53INP1 protein amount was modulated by METTL3 knockdown upon TNF- treatment but not CP treatment, suggesting the existence of a pathway-specific METTL3-TP53INP1 axis. Congruently, METLL3 knockdown sensitized HeLa cells to TNF--mediated apoptosis, which was also validated in a zebrafish larval xenograft model. These results suggest that apoptotic pathway-specific m6A methylation marks exist in cells and TNF--METTL3-TP53INP1 axis modulates TNF--mediated apoptosis in HeLa cells.

cell biology↗

bioRxiv2022

Genomewide m6A mapping uncovers dynamic changes in the m6A epitranscriptome of cisplatin-treated apoptotic HeLa cells

Cisplatin, which is a traditional cancer therapeutic drug, induces apoptosis by modulating a diverse array of gene regulatory mechanisms. However, cisplatin-mediated changes in the m6A methylome is unknown. We employed m6A miCLIP-seq to investigate the effect of m6A methylation events under cisplatin-mediated apoptotic conditions in HeLa cells. Our high-resolution approach revealed numerous m6A marks on 972 target mRNAs with an enrichment on 132 apoptotic mRNAs. Following validation of site-specific m6A events on candidate RNAs, we tracked the fate of candidate mRNAs under METTL3 knockdown and cisplatin treatment conditions. We detected perturbations in the translational efficiency of PMAIP1 and PHLDA1 transcripts based on the polysome profile analyses. Congruently, PMAIP1 and p53 amounts were dependent on METTL3. Additionally, cisplatin-mediated apoptosis was sensitized by METTL3 knockdown. These results suggest that apoptotic pathways are modulated by m6A methylation events and METTL3-p53-PMAIP1 axis modulates cisplatin-mediated apoptosis in HeLa cells.

cell biology↗

bioRxiv2020

Cisplatin-inducible death receptor 5 antisense long non-coding RNA modulates cell cycle and proliferation in HeLa cells

Cisplatin is a chemotherapeutic drug with pleiotropic effects known to modulate the expression of long non-coding RNAs (lncRNAs). With the annotation of many novel lncRNAs, it is imperative to employ a more comprehensive approach to identify cisplatin-mediated changes in the expression of lncRNAs. Next generation sequencing (NGS)-based profiling of total RNAs from cisplatin-treated HeLa cells identified 3489 expressed lncRNAs, of which 1930 and 1051 were up- and downregulated upon cisplatin treatment, respectively. For functional analyses, we selected one of the cisplatin inducible lncRNAs situated antisense to the death receptor 5 and thus named death receptor 5 antisense lncRNA (DR5-AS). Knock-down of DR5-AS lncRNA caused a morphological change in cell shape without inducing any cell death. A second round of NGS-based profiling of total RNAs from DR5-AS-silenced cells revealed differential expression of genes associated with the immune system and cell cycle. Further analyses showed that DR5-AS reduces cell proliferation and causes a cell cycle arrest at S and G2/M phases. These results suggest that cisplatin-mediated reduction in cell proliferation and cell cycle may be mediated by long non-coding RNAs. SignificanceCisplatin is known to induce DNA-damage-induced cell death, which is used in combination chemotherapies in various cancer types. However, many patients develop resistance to cisplatin, which involves both protein-coding and noncoding genes. Although a number of long noncoding RNAs are linked to cisplatin resistance, a more comprehensive study is required. In this study, we took advantage of next-generation-sequencing based lncRNA profiling to unveil the extent of cisplatin inducible lncRNAs in HeLa cells. Additionally, we functionally characterized one of the cisplatin-inducible lncRNA, death receptor 5 antisense. Interestingly, this spesific lncRNA modulates cell morphology, proliferation and cell cycle without affecting cell death.

cell biology↗

bioRxiv2017

Deep sequencing reveals two Jurkat subpopulations with distinct miRNA profiles during camptothecin-induced apoptosis

microRNAs (miRNAs) are small non-coding RNAs of about 19-25 nt that regulate gene expression post-transcriptionally under various cellular conditions, including apoptosis. The miRNAs involved in modulation of apoptotic events in T cells are partially known. However, heterogeneity associated with cell lines makes it difficult to interpret gene expression signatures especially in cancer-related cell lines. Treatment of Jurkat T cell leukemia cell line with the universal apoptotic drug, camptothecin, resulted in identification of two Jurkat sub-populations: one that is sensitive to camptothecin and the other being rather intrinsically resistant. We sorted apoptotic Jurkat cells from the non-apoptotic ones prior to profiling miRNAs through deep sequencing. Our data showed that a total of 184 miRNAs were dysregulated. Interestingly, apoptotic and non-apoptotic sub-populations exhibited a distinct miRNA expression profile. In particular, 6 miRNAs were inversely expressed in these two sub-populations. The pyrosequencing results were validated by real time qPCR. Altogether these results suggest that miRNAs modulate apoptotic events in T cells and that cellular heterogeneity requires careful interpretation of miRNA expression profiles obtained from drug-treated cell lines.

molecular biology↗

bioRxiv2017

Re-arrangements in the cytoplasmic distribution of small RNAs following the maternal-to-zygotic transition in Drosophila embryos

Small RNAs are known to regulate gene expression during early development. However, the dynamics of interaction between small RNAs and polysomes during this process is largely unknown. 0-1h and 7-8h Drosophila embryos were fractionated on sucrose density gradients into four fractions based on A254 reading (1) translationally inactive messengerribonucleoprotein (mRNP); (2) 60S; (3) monosome; and (4) polysome. Comparative analysis of deep-sequencing reads from fractionated and un-fractionated 0-1h and 8-h embryos revealed development-specific co-sedimentation pattern of small RNAs with the cellular translation machinery. Although most miRNAs did not have a specific preference for any state of the translational machinery, we detected fraction-specific enrichment of some miRNAs such as miR-1-3p, -184-39, 5-5p and 263-5p. More interestingly, we observed dysregulation of a subset of miRNAs in fractionated embryos despite no measurable difference in their amount in unfractionated embryos. Transposon-derived endosiRNAs are over-expressed in 7-8h embryos and are associated mainly with the mRNP fraction. However, transposon-derived piRNAs, which are more abundant in 0-1h embryos, co-sediment primarily with the polysome fractions. These results suggest that there appears to be a complex interplay among the small RNAs with respect to their polysome-cosedimention pattern during early development in Drosophila.

developmental biology↗