Understanding how eukaryotic cells assemble their chromatin is a significant research subject in part because several human pathologies including cancer are associated with defects in chromatin assembly. Transporting of newly synthesized histones H3/H4 occurs in a stepwise fashion and is regulated by a variety of protein factors including histone chaperons. In the budding yeast Saccharomyces cerevisiae, the Hif1 protein is an evolutionarily conserved H3/H4-specific histone chaperone and a member of the nuclear Hat1 complex that catalyzes the deposition-related acetylation of newly synthesized histones H4. Hif1 as well as its human homolog NASP have been implicated in an array of chromatin-related processes including histone H3/H4 transport, chromatin assembly, DNA repair and telomeric silencing. In this study, we elucidate structural and functional aspects of Hif1. Through targeted mutational analysis, we demonstrate that the acidic region of yeast Hif1 which interrupts the TPR2 is essential for physical interaction with the Hat1-complex. We also demonstrate that Hif1 requires its C-terminal basic patch for nuclear localization. Furthermore, we provide evidence for the involvement of Hif1 in regulation of histone metabolism by showing that cells lacking HIF1 are both hypersensitive to histone H3 overexpression. Finally, we describe a functional link with a transcriptional regulatory protein Spt2 possibly linking Hif1 to transcription-associated chromatin reassembly. Taken together, our results provide novel mechanistic insights into Hif1 functions and establish it as a key player in various chromatin-associated processes.