Cav1.2 and Cav1.3 L-type calcium channels independently control short- and long-term sensitization to pain.

Key Points: L-type calcium channels in the CNS exist as two subunit forming channels, Cav1.2 and Cav1.3, which are involved in short- and long-term plasticity.

The role of endocytosis on the uptake kinetics of luciferin-conjugated cell-penetrating peptides.

Cell-penetrating peptides (CPPs) are short cationic/amphipathic peptides that can be used to deliver a variety of cargos into cells. However, it is still debated which routes CPPs employ to gain access to intracellular compartments. To assess this, most previously conducted studies have relied on information which is gained by using fluorescently labeled CPPs.

PAIR technology: exon-specific RNA-binding protein isolation in live cells.

RNA-binding proteins (RBPs) are fundamental regulatory proteins for all forms of transcriptional and posttranscriptional control of gene expression. However, isolating RBPs is technically challenging for investigators. Currently, the most widely used techniques to isolate RBPs are in vitro biochemical approaches.

Cellular internalization kinetics of (luciferin-)cell-penetrating peptide conjugates.

Cell-penetrating peptides (CPPs) belong to a class of delivery vectors that have been extensively used for the cellular delivery of various, otherwise impermeable, macromolecules. However, results on the cellular internalization efficacy of CPPs obtained from various laboratories are sometimes challenging to compare because of differences in the experimental setups. Here, for the first time, the cellular uptake kinetics of eight well-established CPPs is compared in HeLa pLuc 705 cells using a recently published releasable luciferin assay.

Secondary structure of cell-penetrating peptides controls membrane interaction and insertion.

The clinical use of efficient therapeutic agents is often limited by the poor permeability of the biological membranes. In order to enhance their cell delivery, short amphipathic peptides called cell-penetrating peptides (CPPs) have been intensively developed for the last two decades. CPPs are based either on protein transduction domains, model peptide or chimeric constructs and have been used to deliver cargoes into cells through either covalent or non-covalent strategies.

Assessing the uptake kinetics and internalization mechanisms of cell-penetrating peptides using a quenched fluorescence assay.

Cell-penetrating peptides (CPPs) have shown great potency for cargo delivery both in vitro and in vivo. Different biologically relevant molecules need to be delivered into appropriate cellular compartments in order to be active, for instance certain drugs/molecules, e.g.

A protocol for PAIR: PNA-assisted identification of RNA binding proteins in living cells.

All aspects of RNA metabolism are regulated by RNA-binding proteins (RBPs) that directly associate with the RNA. Some aspects of RNA biology such as RNA abundance can be readily assessed using standard hybridization technologies. However, identification of RBPs that specifically associate with selected RNAs has been more difficult, particularly when attempting to assess this in live cells.

Immunoprecipitation of mRNA-protein complexes.

Immunoprecipitation of mRNA-protein complexes is a method that can be used to study RNA binding protein (RBP)-RNA interactions. In this protocol, an antibody targeting an RBP of interest is used to immunoprecipitate the RBP and any interacting molecules from a cell lysate. Reverse transcription followed by PCR is then used to identify individual mRNAs isolated with the RBP.

In vivo identification of ribonucleoprotein-RNA interactions.

To understand the role of RNA-binding proteins (RBPs) in the regulation of gene expression, methods are needed for the in vivo identification of RNA-protein interactions. We have developed the peptide nucleic acid (PNA)-assisted identification of RBP technology to enable the identification of proteins that complex with a target RNA in vivo. Specific regions of the 3' and 5' UTRs of ankylosis mRNA were targeted by antisense PNAs transported into cortical neurons by the cell-penetrating peptide transportan 10.

Overcoming methotrexate resistance in breast cancer tumour cells by the use of a new cell-penetrating peptide.

Resistance to chemotherapy limits the effectiveness of anti-cancer drug treatment. Here, we present a new approach to overcome the setback of drug resistance by designing a conjugate of a cell-penetrating peptide and the cytostatic agent methotrexate (MTX). Two different peptides, YTA2 and YTA4, were designed and their intracellular delivery efficiency was characterized by fluorescence microscopy and quantified by fluorometry.

Cell-penetrating peptides: a comparative membrane toxicity study.

Cell-penetrating peptides (CPPs) constitute a new class of delivery vectors with high pharmaceutical potential. However, the abilities of these peptides to translocate through cell membranes can be accompanied by toxic effects resulting from membrane perturbation at higher peptide concentrations. Therefore, we investigated membrane toxicity of five peptides with well-documented cell-penetrating properties, pAntp(43-58), pTAT(48-60), pVEC(615-632), model amphipathic peptide (MAP), and transportan 10, on two human cancer cell lines, K562 (erythroleukemia) and MDA-MB-231 (breast cancer), as well as on immortalized aortic endothelial cells.