The SRB assay has been used since its development in 1990 (Skehan et al., 1990) to inexpensively conduct various screening assays to investigate cytotoxicity in cell based studies (Vichai and Kirtikara, 2006). This method relies on the property of SRB, which binds stoichiometrically to proteins under mild acidic conditions and then can be extracted using basic conditions; thus, the amount of bound dye can be used as a proxy for cell mass, which can then be extrapolated to measure cell proliferation.The protocol can be divided into four main steps: preparation of treatment, incubation of cells with treatment of choice, cell fixation and SRB staining, and absorbance measurement. This assay is limited to manual or semiautomatic screening, and can be used in an efficient and sensitive manner to test chemotherapeutic drugs or small molecules in adherent cells. It also has applications in evaluating the effects of gene expression modulation (knockdown, gene expression upregulation), as well as to study the effects of miRNA replacement on cell proliferation (Kasinski et al., 2015). Bio-protocol is an online peer-reviewed protocol journal. Its mission is to make life science research more efficient and reproducible by curating and hosting high quality, free access protocols.

TRNAs are key adaptor molecules that decipher the genetic code during translation of mRNAs in protein synthesis. In contrast to the traditional view of tRNAs as ubiquitously expressed housekeeping molecules, awareness is now growing that tRNA-encoding genes display tissue-specific and cell type-specific patterns of expression, and that tRNA gene expression and function are both dynamically regulated by post-transcriptional RNA modifications. Moreover, dysregulation of tRNAs, mediated by alterations in either their abundance or function, can have deleterious consequences that contribute to several distinct human diseases, including neurological disorders and cancer. Accumulating evidence shows that reprogramming of mRNA translation through altered tRNA activity can drive pathological processes in a codon-dependent manner. This Review considers the emerging evidence in support of the precise control of functional tRNA levels as an important regulatory mechanism that coordinates mRNA translation and protein expression in physiological cell homeostasis, and highlights key examples of human diseases that are linked directly to tRNA dysregulation.

Targeted cancer therapies, although often effective, have limited utility owing to preexisting primary or acquired secondary resistance. Consequently, agents are sometimes used in combination to simultaneously affect multiple targets. MicroRNA mimics are excellent therapeutic candidates because of their ability to repress multiple oncogenic pathways at once. Here we treated the aggressive Kras;p53 non-small cell lung cancer mouse model and demonstrated efficacy with a combination of two tumor-suppressive microRNAs (miRNAs). Systemic nanodelivery of miR-34 and let-7 suppressed tumor growth leading to survival advantage. This combinatorial miRNA therapeutic approach engages numerous components of tumor cell-addictive pathways and highlights the ability to deliver multiple miRNAs in a safe and effective manner to target lung tissue.

Recent progress in microRNA (miRNA) therapeutics has been strongly dependent on multiple seminal discoveries in the area of miRNA biology during the past two decades. In this review, we focus on the historical discoveries that collectively led to transitioning miRNAs into the clinic. We highlight the pivotal studies that identified the first miRNAs in Caenorhabditis elegans to the more recent reports that have fueled the quest to understand the use of miRNAs as markers for cancer diagnosis and prognosis. In addition, we provide insights as to how unraveling basic miRNA biology has provided a solid foundation for advancing miRNAs, such as miR-34a, therapeutically. We conclude with a brief examination of the current challenges that still need to be addressed to accelerate the path of miRNAs to the clinic: including delivery vehicles, miRNA- and delivery-associated toxicity, dosage, and off target effects.