RNA-seq and CUT&Tag-seq datasets for MDA-MB-231 and MCF-10A cells from: Sp1 mechanotransduction regulates breast cancer cell invasion in engineered viscoelastic extracellular matrices
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Breast cancer progression involves extensive remodeling of the extracellular matrix (ECM), including increased stiffness, altered viscoelasticity (stress relaxation), and elevated collagen levels. While in vitro experiments have revealed a role for each of these factors in individually promoting malignant behavior, their combined effects remain unclear. Here, we engineered alginate-collagen hydrogels with independently tunable stiffness, stress relaxation, and collagen density to dissect how the complex ECM environment regulates cancer cell phenotype. We show that high stiffness, fast stress relaxation, and high collagen density led to changes in cell morphology, marked by decreased roundness, and promoted spheroid invasion in both breast cancer and non-transformed mammary epithelial cells. Single cell migration speed and displacement were greatest in matrices of high stiffness, low collagen density, and slow stress relaxation. RNA-seq and Cleavage Under Targets and Tagmentation (CUT&am..., For RNA-seq, cells were extracted from alginate-collagen matrices by rocking them in Falcon tubes containing 2.5 mg/ml collagenase (Sigma, Cat#C0130) solution in PBS for 30 minutes at 37°C. After this, the tubes were centrifuged, and the supernatant was removed. Following this, the pellet was dissolved in 10 ml ice-cold EDTA (50 mM) and placed on a rotator for 10 mins. The tubes were centrifuged, and the supernatant was then removed. The cell pellet was then lysed using Trizol (Life Technologies), and RNA was extracted using the total RNA mini prep kit according to the manufacturerâs instructions (Epoch Life Sciences). Bulk-mRNA-sequencing library prep was done using the Cel-Seq2 pipeline as described previously. Briefly, 10 ng RNA was reverse-transcribed using the CelSeq2 RT-primer, DTT (0.1 M), dNTPs (New England Biolabs, Cat# N0447l), and Superscript II reverse transcriptase (Invitrogen, Cat# 18064014), followed by second-strand synthesis using RNAseH (ThermoFisher Scientific, Cat# E..., # RNA-seq and CUT&Tag-seq datasets for MDA-MB-231 and MCF-10A cells encapsulated in alginate-collagen viscoelastic matrices. Dataset DOI: [10.5061/dryad.dfn2z35fm](https://doi.org/10.5061/dryad.dfn2z35fm) ## Description of the data and file structure RNA-seq of MDA-MB-231 and MCF-10A cells encapsulated in Soft/Stiff; Slow relaxing/Fast relaxing, low collagen/high collagen density alginate-based hydrogels. 3 replicates per mechanical condition for MDA-MB-231 cells and 2 replicates per mechanical condition for MCF-10A cells. r1 , r2, r3 are replicate numbers. CUT&Tag seq of Sp1 in MDA-MB-231 cells encapsulated in Soft/Stiff; Slow relaxing/Fast relaxing, low collagen density alginate-based hydrogels. 2 replicates per mechanical condition. ### Files and variables #### File: cut-n-tag_metadata_file.xlsx **Description:** Metadata file containing sample names, respective fastq files, and .bw files for CUT&Tag-seq dataset. #### File: rna-seq_metadata_file.xlsx **Description:** Metadat...,
乳腺癌进展涉及细胞外基质（extracellular matrix, ECM）的广泛重塑，包括基质硬度增加、粘弹性改变（应力松弛）以及胶原蛋白水平升高。尽管体外实验已证实上述各因素均可单独促进恶性表型，但它们的联合效应仍未明确。本研究构建了可独立调控硬度、应力松弛及胶原蛋白密度的海藻酸-胶原蛋白水凝胶，以解析复杂ECM微环境对癌细胞表型的调控机制。研究发现，高硬度、快速应力松弛及高胶原蛋白密度可改变细胞形态，表现为细胞圆度降低，并可促进乳腺癌细胞与非转化乳腺上皮细胞的球状体侵袭；而单细胞迁移速度和位移在高硬度、低胶原蛋白密度、慢应力松弛的基质中最为显著。 关于RNA-seq实验：将包裹于海藻酸-胶原蛋白基质中的细胞置于含2.5 mg/ml胶原酶（Sigma，货号C0130）的PBS溶液的Falcon管中，于37℃摇育30分钟以解离细胞。随后离心并移除上清液，将沉淀重悬于10 ml冰浴EDTA（50 mM）中，于旋转仪上孵育10分钟，再次离心并移除上清液。之后使用Trizol（Life Technologies）裂解细胞沉淀，并按照制造商操作手册（Epoch Life Sciences）使用总RNA迷你提取试剂盒提取总RNA。 批量mRNA测序文库制备采用此前报道的Cel-Seq2流程：简要而言，使用CelSeq2 RT引物、DTT（0.1 M）、dNTPs（New England Biolabs，货号N0447l）及Superscript II逆转录酶（Invitrogen，货号18064014）对10 ng RNA进行逆转录，随后使用RNAseH（ThermoFisher Scientific，货号E...）进行第二链合成。 本数据集包含包裹于海藻酸-胶原蛋白粘弹性基质中的MDA-MB-231与MCF-10A细胞的RNA-seq及CUT&Tag-seq数据。数据集DOI：[10.5061/dryad.dfn2z35fm](https://doi.org/10.5061/dryad.dfn2z35fm) ### 数据与文件结构说明 1. RNA-seq数据：对包裹于软/硬、慢松弛/快松弛、低/高胶原蛋白密度海藻酸基质中的MDA-MB-231和MCF-10A细胞进行RNA测序。其中MDA-MB-231细胞每个力学条件设置3个生物学重复，MCF-10A细胞每个力学条件设置2个生物学重复，r1、r2、r3为重复编号。 2. CUT&Tag测序数据：对包裹于软/硬、慢松弛/快松弛、低胶原蛋白密度海藻酸基质中的MDA-MB-231细胞进行Sp1的CUT&Tag测序，每个力学条件设置2个生物学重复。 #### 文件：cut-n-tag_metadata_file.xlsx **描述：** 该元数据文件包含CUT&Tag-seq数据集的样本名称、对应fastq文件及.bw文件。 #### 文件：rna-seq_metadata_file.xlsx **描述：** 该元数据文件包含RNA-seq数据集的相关元数据...
