Biology subjects

Aleksic, J.

Publications and source records attributed to Aleksic, J..

4 recordsLinked to original sources

bioRxiv2024

Long-read sequencing to detect full-length protein-protein interactions

Given the increased predictions on interactome size and demand for protein function information, methods for detecting protein-protein interactions remain a significant development area. The all-vs.-all sequencing (AVA-Seq) method utilizes a convergent fusion plasmid design to make two-hybrid technology amenable to next-generation sequencing. Here, we further innovate to take advantage of synthetic DNA technologies and Oxford Nanopore Technologies long-read sequencing improvements to allow us to determine full-length protein-protein interactions. Here, using this approach we recovered 159 protein-protein interactions from a set of 57 human proteins using multiple forms of validation. Further, when referencing a human gold standard set of interactions, eight full-length protein-protein interactions were recovered from an expected 28 interaction pairs (28.6%), a typical recovery rate for two-hybrid technologies. The AVA-Seq, in combination with the ease of synthetic DNA production and the MinION platform, offers a low-cost, high-throughput alternative for determining protein-protein interactions, which can be utilized in research labs at all stages. 3 Key PointsO_LIFirst application of long-read sequencing for full-length protein-protein interaction studies. C_LIO_LIThe recovery rate of the AVA-Seq method using full-length proteins is on par with other leading methods. C_LIO_LIAdvances in synthetic biology and sequencing technologies make full-length protein interactomes affordable and accessible. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/586447v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@1d81412org.highwire.dtl.DTLVardef@10dbb19org.highwire.dtl.DTLVardef@76a04borg.highwire.dtl.DTLVardef@15baf0_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2022

Identifying novel interactions of the colon-cancer related APC protein with Wnt-pathway nuclear transcription factors

BackgroundColon cancer is often driven by mutations of the adenomatous polyposis coli (APC) gene, an essential tumor suppressor gene of the Wnt {beta}-catenin signaling pathway. APC and its interactions in the cytoplasm have been well studied, however various groups have also observed its presence in the nucleus. Identifying novel interactions of APC in the Wnt pathway will provide an opportunity to better understand the nuclear role of APC and ultimately identify potential cancer treatment targets. MethodsWe used the all-vs-all sequencing (AVA-Seq) method to interrogate the interactome of protein fragments spanning most of the 60 Wnt {beta}-catenin pathway proteins. Using protein fragments identified the interacting regions between the proteins with more resolution than a full-length protein approach. Pull-down assays were used to validate a subset of these interactions. Results74 known and 703 novel Wnt {beta}-catenin pathway protein-protein interactions were recovered in this study. There were 8 known and 31 novel APC protein-protein interactions. Novel interactions of APC and nuclear transcription factors TCF7, JUN, FOSL1, and SOX17 were particularly interesting and confirmed in validation assays. ConclusionBased on our findings of novel interactions between APC and transcription factors and previous evidence of APC localizing to the nucleus, we suggest APC may compete and repress CTNNB1. This would occur through the binding of the transcription factors (JUN, FOSL1, TCF7) to regulate the Wnt signaling pathway including through enhanced marking of CTNNB1 for degradation in the nucleus by APC binding with SOX17. Additional novel Wnt {beta}-catenin pathway protein-protein interactions from this study could lead researchers to novel drug designs for cancer.

molecular biology↗

bioRxiv2021

High-resolution protein fragment interactions using AVA-Seq on a human reference set

Protein-protein interactions (PPIs) are important in understanding numerous aspects of protein function. Here, the recently developed all-vs-all sequencing (AVA-Seq) approach to determine protein-protein interactions was tested on a gold-standard human protein interaction set (hsPRS-v2). Initially, these data were interpreted strictly from a binary PPI perspective to compare AVA-Seq to other binary PPI methods tested on the same hsPRS-v2. AVA-Seq recovered 20 of 47 (43%) binary PPIs from this reference set comparing favorably with other methods. The same experimental data allowed for the determination of >500 known and novel PPIs including interactions between wildtype fragments of tumor protein p53 and minichromosomal maintenance complex proteins 2, and 5 (MCM2 and MCM5) that could be of interest in human disease. Additional results gave a better understanding of why interactions might be missed using AVA-Seq and aide future PPI experimental design for maximum recovery of information.

genomics↗

bioRxiv2018

Genomic data provides new insights on the demographic history and the extent of recent material transfers in Norway spruce

Primeval forests are today exceedingly rare in Europe and transfer of forest reproductive material for afforestation and improvement have been very common, especially over the last two centuries. This can be a serious impediment when inferring past population movements in response to past climate changes such as the last glacial maximum (LGM), some 18,000 years ago. In the present study, we genotyped 1,672 individuals from three Picea species (P. abies, P. obovata, and P. omorika) at 400K SNPs using exome capture to infer the past demographic history of Norway spruce and estimate the amount of recent introduction used to establish the Norway spruce breeding program in Southern Sweden. Most of these trees belong to P. abies and originate from the base population of the Swedish breeding program. Others originate from populations across the natural ranges of the three species. Of the 1,499 individuals stemming from the breeding program, a large proportion corresponds to recent introductions. The split of P. omorika occurred 23 million years ago (mya), while the divergence between P. obovata and P. abies began 17.6 mya. Demographic inferences retrieved the same main clusters within P. abies than previous studies, i.e. a vast northern domain ranging from Norway to central Russia, where the species is progressively replaced by Siberian spruce (P. obovata) and two smaller domains, an Alpine domain, and a Carpathian one, but also revealed further subdivision and gene flow among clusters. The three main domains divergence was ancient (15 mya) and all three went through a bottleneck corresponding to the LGM. Approximately 17% of P. abies Nordic domain migrated from P. obovata ~103K years ago, when both species had much larger effective population sizes. Our analysis of genome-wide polymorphism data thus revealed the complex demographic history of Picea genus in Western Europe and highlighted the importance of material transfer in Swedish breeding program.

evolutionary biology↗