Despite extensive in vitro studies identifying a myriad of cellular transcription factors that bind the human immunodeficiency virus type 1 5' long terminal repeat (LTR), the relative contribution of these factors to human immunodeficiency virus type 1 replication in infected individuals remains obscure. To address this question, we investigated 478 proviral quasispecies derived from uncultured peripheral blood mononuclear cells of 42 patients representing all stages of infection. In addition to highly conserved TATA box, SP-1, and NF-kappaB sites, the Ets core and an adjacent 5'-ACYGCTGA-3' motif were extremely conserved. Importantly, the most frequent naturally occurring length polymorphism (MFNLP) duplicated 5'-ACYGCTGA-3' motifs in LTRs in which this same motif was disrupted or in LTRs in which a single point mutation to the Ets core ablated binding of c-Ets 1 and another factor distinct from both c-Ets 1 and Elf 1. The MFNLP's location was precise (position -121) and surprisingly frequent (38% of patients) and demarcated LTR Nef-coding sequences from LTR noncoding sequences that appear to be evolving independently. Aside from these features, we found no definitive clinical or transcription phenotype common to all MFNLP LTRs. We also found previously described and novel point polymorphisms, including some conferring TAR-dependent and TAR- independent Tat unresponsiveness, and showed that differential binding of nuclear factor(s) to a TCTAA TATA box variant may be the mechanism for the latter.