Proteins that emerge de novo from noncoding DNA could negatively or positively influence cellular physiology in the sense of providing a possible adaptive advantage. Here, we employ two approaches to study such effects in a human cell line by expressing random sequences and mouse de novo genes that lack homologs in the human genome. We show that both approaches lead to differential growth effects of the cell clones dependent on the sequences they express. For the random sequences, 53% of the clones decreased in frequency, and about 8% increased in frequency in a joint growth experiment. Of the 14 mouse de novo genes tested in a similar joint growth experiment, 10 decreased, and 3 increased in frequency. When individually analysed, each mouse de novo gene triggers a unique transcriptomic response in the human cells, indicating mostly specific rather than generalized effects. Structural analysis of the de novo gene open reading frames (ORFs) reveals a range of intrinsic disorder scores and/or foldability into alpha-helices or beta sheets, but these do not correlate with their effects on the growth of the cells. Our results indicate that de novo evolved ORFs could easily become integrated into cellular regulatory pathways, since most interact with components of these pathways and could therefore become directly subject to positive selection if the general conditions allow this.