The endoplasmic reticulum (ER) serves as a central hub for protein production and sorting in eukaryotic cells, processing approximately one-third of the cellular proteome. Protein targeting to the ER occurs through multiple pathways that operate both during and independent of translation. The classical translation-dependent pathway, mediated by cytosolic factors like signal recognition particle, recognizes signal peptides or transmembrane helices in nascent proteins, while translation-independent mechanisms utilize RNA-based targeting through specific sequence elements and RNA-binding proteins. At the core of these processes lies the Sec61 complex, which undergoes dynamic conformational changes and coordinates with numerous accessory factors to facilitate protein translocation and membrane insertion across and into the endoplasmic reticulum membrane. This review focuses on the molecular mechanisms of protein targeting to the ER, from the initial recognition of targeting signals to the dynamics of the translocation machinery, highlighting recent discoveries that have revealed unprecedented complexity in these cellular trafficking pathways.