Evidence for a base-paired region of hepatitis B virus pregenome encapsidation signal which influences the patterns of precore mutations abolishing HBe protein expression

Abstract

In two natural HBe-minus hepatitis B virus mutants, expression of HBe protein was abrogated by a nonsense mutation at precore codon 28 and a frameshift mutation at codon 29, respectively. Both mutants contained an additional nucleotide substitution(s) which was found by transfection experiments to be required for efficient packaging of pregenomic RNA. The observed mutational profiles were consistent with the presence of a base-paired region of the pregenome encapsidation signal overlapping the HBe-coding sequence. Results obtained with artificial mutants with significant changes in the primary sequence suggested that base pairing is required but insufficient for efficient pregenome packaging. However, the predicted first four base pairs of the stem are dispensable.

Structure and function of the encapsidation signal of hepadnaviridae

“…Mutations in the lower stem. Base pairing and sequencespecific determinants (when more than four nts are altered) in the upper part of the lower stem are important for ε to function in encapsidation [18,29,63,64], initiation of reverse transcription in HBV [29] and activation of POL in DHBV [25,43]. This was confirmed by the observation that interaction of POL-ε was optimal between POL and the cognate lower stem of ε [25].…”

“…Disruption of lower stem base pairs near the bulge affect the function of ε more severely [19] than disruption further down. In fact, the four lowest nucleotides are dispensable for encapsidation of pgRNA [64] and in WHV ε they are not base paired [5,22].…”

Predictive value of precore hepatitis B virus mutations in spontaneous and interferon-induced hepatitis B e antigen clearance

“…Elevated pretreatment ALT level has been identified as a predictive factor for favorable response to IFN therapy, perhaps 2). 14,[17][18][19] reflecting endogenous immune lysis of infected hepatocytes.1"21 '22 We and others have noted that M2 (A-1896) tends to emerge or become predominant around the time of ALT flare.*."' Thus, it seems that in patients with protracted or incomplete immune clearance, mutants that are capable of evading immune surveillance or replicating a t higher levels may develop or are selected.…”

Clinical implications of mutations in the hepatitis B virus genome

“…3). The presence of a C at position 1858 already creates a base pairing with the G at position 1896, therefore precluding the G → A mutation [55][56][57][58] unless a C → T mutation at 1858 is also present [54,57].…”

“…Point mutations leading to failure to initiate (first codon [59,60]) or to premature termination of the HBeAg translation at the second codon [61], as well as frameshifts due to deletions or insertions [55,60,[62][63][64], account for approximately 10% of the HBV population with the HBe-minus phenotype [65].…”