Nagoya University · 생화학·유전·분자생물학
Kohei Nishimura 교수의 연구실은 단백질 기능 연구를 위한 고감도 조절 탈형성 기술, 특히 auxin-inducible degron(AID) 시스템의 개발과 응용에 초점을 맞추고 있습니다. 유전자 조작이 가능한 효모와 동물세포를 대상으로, 저농도의 합성 유도제로도 빠르고 정밀하게 목표 단백질을 제거할 수 있는 슈퍼민감도 AID 시스템을 개발하여, 생물학적 기능 분석과 약물 타겟 발굴에 기여하고 있습니다. 특히, 단일 도메인 항체를 활용한 AlissAID 시스템과 고친화성 TIR1 변형체를 통해 세포 내 단백질 조절의 정밀도와 유연성을 극대화하고 있습니다.
표시된 성과는 수집된 데이터 기준으로 산출되며, 일부 차이가 있을 수 있습니다.
The auxin-inducible degron (AID) system allows the rapid and reversible proteolysis of proteins of interest, and enables the generation of conditional mutants of budding yeast. The construction of budding yeast AID mutants is simple, and the effect of depletion of essential proteins on proliferation can be confirmed by analyzing their phenotype. In this protocol, we describe a procedure to generate AID mutants of budding yeast via a simple transformation using PCR-amplified DNA. We also describe
The auxin-inducible degron (AID) system enables rapid depletion of target proteins within the cell by applying the natural auxin IAA. The AID system is useful for investigating the physiological functions of essential proteins; however, this system generally requires high dose of auxin to achieve effective depletion in vertebrate cells. Here, we describe a super-sensitive AID system that incorporates the synthetic auxin derivative 5-Ad-IAA and its high-affinity-binding partner OsTIR1F74A. The su
The centromere is an important genomic locus for chromosomal segregation. Although the centromere is specified by sequence-independent epigenetic mechanisms in most organisms, it is usually composed of highly repetitive sequences, which associate with heterochromatin. We have previously generated various chicken DT40 cell lines containing differently positioned neocentromeres, which do not contain repetitive sequences and do not associate with heterochromatin. In this study, we performed systema
Abstract A uxin- I nducible D egron (AID) technology enables conditional depletion of targeted proteins. However, the applicability of the AID in vertebrate cells has been limited due to cytotoxicity caused by high auxin concentrations. Here, we establish an improved AID system using an engineered orthogonal auxin-TIR1 pair, which exhibits over 1,000 times stronger binding. With ~1,000-fold less auxin concentration, we achieved to generate the AID-based knockout cells in various human and mouse
Conditional control of target proteins using the auxin-inducible degron (AID) system provides a powerful tool for investigating protein function in eukaryotes. Here, we established an Affinity-linker based super-sensitive auxin-inducible degron (AlissAID) system in budding yeast by using a single domain antibody (a nanobody). In this system, target proteins fused with GFP or mCherry were degraded depending on a synthetic auxin, 5-Adamantyl-IAA (5-Ad-IAA). In AlissAID system, nanomolar concentrat
This review explores various methods for modulating protein stability to achieve target protein degradation, which is a crucial aspect in the study of biological processes and drug design. Thirty years have passed since the introduction of heat-inducible degron cells utilizing the N-end rule, and methods for controlling protein stability using the ubiquitin-proteasome system have moved from academia to industry. This review covers protein stability control methods, from the early days to recent
In this article, we have developed the gated CNT-FED fabrication process with the polymer insulator. CNTs which are lying down on the substrate are successfully activated with laser.
Inducing loss of function of a target protein using methods such as gene knockout is a powerful and useful strategy for analyzing protein function in cells. In recent years, the CRISPR/Cas-9-based gene knockout technology has been widely used across a variety of eukaryotes; however, this type of simple gene knockout strategy is not applicable to essential genes, which require a conditional knockout system. The auxin-inducible degron (AID) system enables rapid depletion of the target protein in a
Abstract Protein analysis strategies involving targeted protein degradation are powerful approaches to determine gene functions. Auxin-inducible degron (AID) is among the most widely used methods for target protein knockdown. This system enables the rapid depletion of AID-tagged target proteins in an auxin-dependent manner. Various improved AID methods have been developed to date; however, the requirement to tag the target protein remains a common challenge. Here, we demonstrated the efficiency
