Biology subjects

Hammarskjold, M.-L.

Publications and source records attributed to Hammarskjold, M.-L..

4 recordsLinked to original sources

bioRxiv2025

Identification and Characterization of Functional and Trans-dominant Negative HERV-K (HML-2) Rec Proteins Encoded in the Human Genome.

Human Endogenous Retroviruses K (HERV-K) of the HML-2 subgroup are the most recently integrated and biologically active retroviral elements within the human genome. The HERV-K Rec protein, a functional homolog of HIV Rev and HTLV Rex, is necessary for the nuclear export of viral mRNAs with retained introns. However, the diversity of Rec proteins encoded in the human genome and their functional capacities have remained largely unexplored. We identified full-length Rec protein sequences in the human genome and selected 23 variants for functional characterization. Using a dual-color fluorescent reporter and a complementary ELISA assay, we found that Rec proteins from only 7 genomic loci were functional. In addition, a subset of the non-functional Rec proteins exerted trans-dominant negative effects. Detailed mutational analysis of the most potent inhibitory variant, encoded by the HERV-K provirus 12q14.1, displayed only 2 amino acid changes (H2N and del34E) relative to the prototypical functional Rec protein encoded by the reconstructed consensus HERV-K (HERV-K Con). Insertion of a glutamic acid at position 34 restored the functional activity, while the substitution of histidine with asparagine at position 2 did not. Our results unveil an unexpected complexity in HERV-K (HML-2) post-transcriptional regulation, with Rec variants displaying a spectrum of activities ranging from robust nuclear export of RNA with retained introns to trans-dominant negative inhibition of Rec function. These findings expand our understanding of the regulatory landscape governing HERV-K (HML-2) expression and suggest mechanisms by which different HERV-K Rec proteins may influence host cell biology and pathology, including oncogenesis.

cell biology↗

bioRxiv2025

Rev-RRE activity modulates HIV-1 replication and latency reactivation: Implications for viral persistence and cure strategies

The HIV-1 Rev-RRE regulatory axis plays a crucial role in viral replication by facilitating the nucleo-cytoplasmic export and expression of viral mRNAs with retained introns. In this study, we investigated the impact of variation in Rev-RRE functional activity on HIV-1 replication kinetics and reactivation from latency. Using a novel HIV-1 clone with an interchangeable Rev cassette, we engineered viruses with different Rev functional activities and demonstrated that higher Rev-RRE activity confers greater viral replication capacity while maintaining a constant level of Nef expression. In addition, a low Rev activity virus rapidly acquired a compensatory mutation in the RRE that significantly increased Rev-RRE activity and replication. In a latency model, proviruses with differing Rev-RRE activity levels varied in the efficiency of viral reactivation, affecting both initial viral release and subsequent replication kinetics. These results demonstrate that activity differences in the Rev-RRE axis among different viral isolates have important implications for HIV replication dynamics and persistence. Importantly, our findings indicate that bolstering Rev/RRE activity could be explored as part of latency reversal strategies in HIV cure efforts. Author SummaryThe activity of the HIV-1 Rev-RRE axis is essential for viral replication and varies among primary viral isolates. However, the role of this for viral fitness, evolution, and persistence has not previously been investigated. Our results show that during in vitro replication, there is a selective fitness advantage for viruses with higher Rev-RRE activity and that HIV has the ability to fine tune this regulatory system with minimal sequence changes. Additionally, the maintenance of Nef expression in low Rev activity viruses suggests a potential mechanism for balancing immune evasion and replication capacity in different selection landscapes within a host. We also show that viruses with low Rev-RRE activity are more difficult to reverse from latency than viruses with higher Rev-RRE activity. Thus, differences in provirus Rev-RRE activity may be a barrier to developing effective latency reversal strategies. These findings provide new insights into the complex roles that the Rev/RRE axis plays in functionality, viral fitness, evolution, and persistence.

microbiology↗

bioRxiv2019

An assay to measure the activity of RNA elements and proteins that promote the expression of mRNA with retained introns

The ability to overcome cellular restrictions that exist for the export and translation of mRNAs with retained introns is a requirement for the replication of retroviruses and also for the expression of many mRNA isoforms transcribed from cellular genes. In some cases, RNA structures have been identified in the mRNA that directly interact with cellular factors to promote the export and expression of isoforms with retained introns. In other cases, a viral protein is also required to act as an adapter. In this report we describe a novel vector system that allows measurement of the ability of cis- and trans-acting factors to promote the export and translation of mRNA with retained introns. One reporter vector used in this system is derived from an HIV proviral clone engineered to express two different fluorescent proteins from spliced and unspliced transcripts. The ratio of fluorescent signals is a measurement of the efficiency of export and translation. A second vector utilizes a third fluorescent protein to measure the expression of viral export proteins that interact with some of the export elements. Both vectors can be packaged into viral particles and be used to transduce cells, allowing expression at physiological levels from the integrated vector.

molecular biology↗

bioRxiv2018

Evolution of the HIV-1 RRE during natural infection reveals key nucleotide changes that correlate with altered structure and increased activity over time

AbstractThe HIV-1 Rev Response Element (RRE) is a cis-acting RNA element characterized by multiple stem-loops. Binding and multimerization of the HIV Rev protein on the RRE promotes nucleocytoplasmic export of incompletely spliced mRNAs, an essential step in HIV replication. Most of our understanding of the Rev-RRE regulatory axis comes from studies of lab-adapted HIV clones. However, in human infection, HIV evolves rapidly and mechanistic studies of naturally occurring Rev and RRE sequences are essential to understanding this system. We previously described the functional activity of two RREs found in circulating viruses in a patient followed during the course of HIV infection. The "early" RRE was less functionally active than the "late" RRE despite differing in sequence by only four nucleotides. In this study, we describe the sequence, function, and structural evolution of circulating RREs in this patient using plasma samples collected over six years of untreated infection. RRE sequence diversity varied over the course of infection with evidence of selection pressure that led to sequence convergence as disease progressed. An increase in RRE functional activity was observed over time, and a key mutation was identified that correlates with a major conformational change in the RRE and increased functional activity. Additional mutations were found that may have contributed to increased activity as a result of greater Shannon entropy in RRE stem-loop II, which is key to primary Rev binding. ImportanceHIV-1 replication requires interaction of the viral Rev protein with a cis-acting regulatory RNA, the Rev Response Element (RRE), whose sequence changes over time during infection within a single host. In this study, we show that the RRE is subject to selection pressure and that RREs from later time points in infection tend to have higher functional activity. Differences in RRE functional activity are attributable to specific changes in RNA structure. Our results suggest that RRE evolution during infection may be important for HIV pathogenesis and that efforts to develop therapies acting on this viral pathway should take this into account.

molecular biology↗